Note: Descriptions are shown in the official language in which they were submitted.
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MOLECULES FOR DISEASE DETECTION AND TREATMENT
TECHNICAL FIELD
The invention relates to novel nucleic acids, molecules for disease detection
and treatment
encoded by these nucleic acids, and to the use of these nucleic acids and
proteins in the diagnosis,
treatment, and prevention of cell proliferative, autoimmune/inflammatory,
developmental, and
neurological disorders. The invention also relates to the assessment of the
effects of exogenous
compounds on the expression of nucleic acids and molecules for disease
detection and treatment.
BACKGROUND OF THE INVENTION
It is estimated that only 2% of mammalian DNA encodes proteins, and only a
small fraction
of the genes that encode proteins are actually expressed in a particular cell
at any time. The various
types of cells in a multicellular organism differ dramatically both in
structure and function, and the
identity of a particular cell is conferred by its unique pattern of gene
expression. In addition,
different cell types express overlapping but distinctive sets of genes
throughout development. Cell
growth and proliferation, cell differentiation, the immune response,
apoptosis, and other processes
that contribute to organismal development and survival are governed by
regulation of gene
expression. Appropriate gene regulation also ensures that cells function
efficiently by expressing
only those genes whose functions are required at a given time. Factors that
influence gene expression
include extracellular signals that mediate cell-cell communication and
coordinate the activities of
different cell types. Gene expression is regulated at the level of DNA and RNA
transcription, and at
the level of mRNA translation.
Aberrant expression or mutations in genes and their products may cause, or
increase
susceptibility to, a variety of human diseases such as cancer and other cell
proliferative disorders.
The identification of these genes and their products is the basis of an ever-
expanding effort to find
markers for early detection of diseases and targets for their prevention and
treatment. For example,
cancer represents a type of cell proliferative disorder that affects nearly
every tissue in the body. The
development of cancer, or oncogenesis, is often correlated with the conversion
of a normal gene into
a cancer-causing gene, or oncogene, through abnormal expression or mutation.
Oncoproteins, the
products of oncogenes, include a variety of molecules that influence cell
proliferation, such as growth
factors, growth factor receptors, intracellular signal transducers, nuclear
transcription factors, and
cell-cycle control proteins. In contrast, tumor-suppressor genes are involved
in inhibiting cell
proliferation. Mutations which reduce or abrogate the function of tumor-
suppressor genes result in
aberrant cell proliferation and cancer. Thus a wide variety of genes and their
products have been
found that are associated with cell proliferative disorders such as cancer,
but many more may exist
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that are yet to be discovered.
DNA-based arrays can provide an efficient, high-throughput method to examine
gene
expression and genetic variability. For example, SNPs, or single nucleotide
polymorphisms, are the
most common type of human genetic variation. DNA-based arrays can dramatically
accelerate the
discovery of SNPs in hundreds and even thousands of genes. Likewise, such
arrays can be used for
SNP genotyping in which DNA samples from individuals or populations are
assayed for the presence
of selected SNPs. These approaches will ultimately lead to the systematic
identification of all genetic
variations in the human genome and the correlation of certain genetic
variations with disease
susceptibility, responsiveness to drug treatments, and other medically
relevant information. (See, for
example, Wang, D.G. et al. (1998) Science 280:1077-1082.)
DNA-based array technology is especially important for the rapid analysis of
global gene
expression patterns. For example, genetic predisposition, disease, or
therapeutic treatment may
directly or indirectly affect the expression of a large number of genes in a
given tissue. In this case, it
is useful to develop a profile, or transcript image, of all the genes that are
expressed and the levels at
which they are expressed in that particular tissue. A profile generated from
an individual or
population affected with a certain disease or undergoing a particular therapy
may be compared with a
profile generated from a control individual or population. Such analysis does
not require knowledge
of gene function, as the expression profiles can be subjected to mathematical
analyses which simply
treat each gene as a marker. Furthermore, gene 'expression profiles may help
dissect biological
pathways by identifying all the genes expressed, for example, at a certain
developmental stage, in a
particular tissue, or in response to disease or treatment. (See, for example,
Lander, E.S. et al. (1996)
Science 274:536-539.)
Certain genes are known to be associated with diseases because of their
chromosomal
location, such as the genes in the myotonic dystrophy (DM) regions of mouse
and human. The
mutation underlying DM has been localized to a gene encoding the DM-kinase
protein, but another
active gene, DMR-N9, is in close proximity to the DM-kinase gene (Jansen, G.
et al. ( 1992) Nat.
Genet. 1:261-266). DMR-N9 encodes a 650 amino acid protein that contains WD
repeats, motifs
found in cell signaling proteins. DMR-N9 is expressed in all neural tissues
and in the testis,
suggesting a role for DMR-N9 in the manifestation of mental and testicular
symptoms in severe cases
of DM (Jansen, G. et al. (1995) Hum. Mol. Genet. 4:843-852).
Other types of signaling proteins include the WW domain, which consists of 35-
40 amino
acids and is characterized by four well-conserved aromatic residues, two of
which are tryptophan.
The secondary structure of the WW domain consists of a slightly bent three-
stranded antiparallel
sheet. This domain has been reported in a wide variety of proteins, including
human Pin 1 and Ras
GAP-related protein. The presence of the WW domain in diverse proteins is
involved in signaling,
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regulatory, and cytoskeletal functions. Defects in WW domains containing
proteins are associated
with human diseases such as Liddle Syndrome (Pirozzi, G. et al. (1997) J.
Biol. Chem. 272:14611-
14616). The tetratricopeptide repeat (TPR) is composed of a degenerate 34
amino acid sequence
present in various proteins. Their array as multiple motifs enable formation
of scaffolds that mediate
protein-protein interactions and assembly of multiprotein complexes (Das, A.K.
et al. (1998) EMBO
J. 17:1192-1199). CheB methylesterase catalyzes hydrolysis of receptor
glutamine or
methylglutamate side-chains to glutamic acid, and belongs to a large family of
response regulator
proteins in which N-terminal regulatory domains control the activities of C-
terminal effector domains.
Other genes are identified based upon their expression patterns or association
with disease
syndromes. For example, autoantibodies to subcellular organelles are found in
patients with systemic
rheumatic diseases. A recently identified protein, golgin-67, belongs to a
family of Golgi
autoantigens having alpha-helical coiled-coil domains (Eystathioy, T. et al.
(2000) J. Autoimmun.
14:179-187). The Stac gene was identified as a brain specific, developmentally
regulated gene. The
Stac protein contains an SH3 domain, and is thought to be involved in neuron-
specific signal
transduction (Suzuki, H. et al. (1996) Biochem. Biophys. Res. Commun. 229:902-
909).
Expression profiling
Microarrays are analytical tools used in bioanalysis. A' microarray has a
plurality of
molecules spatially distributed over, and stably associated with, the surface
of a solid support.
Microarrays of polypeptides, polynucleotides, and/or antibodies have been
developed and find use in
a variety of applications, such as gene sequencing, monitoring gene
expression, gene mapping,
bacterial identification, drug discovery, and combinatorial chemistry.
One area in particular in which microarrays find use is in gene expression
analysis. Array
technology can provide a simple way to explore the expression of a single
polymorphic gene or the
expression profile of a large number of related or unrelated genes. When the
expression of a single
gene is examined, arrays are employed to detect the expression of a specific
gene or its variants.
When an expression profile is examined, arrays provide a platform for
identifying genes that are
tissue specific, are affected by a substance being tested in a toxicology
assay, are part of a signaling
cascade, carry out housekeeping functions, or are specifically related to a
particular genetic
predisposition, condition, disease, or disorder.
There is a need in the art for new compositions, including nucleic acids and
proteins, for the
diagnosis, prevention, and treatment of cell proliferative,
autoimmune/inflammatory, developmental,
and neurological disorders.
SUMMARY OF THE INVENTION
Various embodiments of the invention provide purified polypeptides, molecules
for disease
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detection and treatment, referred to collectively as 'MDDT' and individually
as 'MDDT-1,' 'MDDT-
2,' 'MDDT-3,' 'MDDT-4,' 'MDDT-5,' 'MDDT-6,' 'MDDT-7,' 'MDDT-8,' 'MDDT-9,'
'MDDT-
10,' 'MDDT-11,' 'MDDT-12,' 'MDDT-13,' 'MDDT-14,' 'MDDT-15,' 'MDDT-16,' 'MDDT-
17,'
'MDDT-18,' 'MDDT-19,' 'MDDT-20,' 'MDDT-21,' 'MDDT-22,' 'MDDT-23,' 'MDDT-24,'
'MDDT-25,' 'MDDT-26,' 'MDDT-27,' 'MDDT-28,' 'MDDT-29,' 'MDDT-30,' 'MDDT-31,'
'MDDT-32,' 'MDDT-33,' 'MDDT-34,' 'MDDT-35,' 'MDDT-36,' 'MDDT-37,' 'MDDT-38,'
'MDDT-39,' 'MDDT-40,' 'MDDT-41,' 'MDDT-42,' 'MDDT-43,' 'MDDT-44,' 'MDDT-45,'
'MDDT-46,' 'MDDT-47,' and 'MDDT-48' and methods for using these proteins and
their encoding
polynucleotides for the detection, diagnosis, and treatment of diseases and
medical conditions.
Embodiments also provide methods for utilizing the purified molecules for
disease detection and
treatment and/or their encoding polynucleotides for facilitating the drug
discovery process, including
determination of efficacy, dosage, toxicity, and pharmacology. Related
embodiments provide
methods for utilizing the purified molecules for disease detection and
treatment and/or their encoding
polynucleotides for investigating the pathogenesis of diseases and medical
conditions.
An embodiment provides an isolated polypeptide selected from the group
consisting of a) a
polypeptide comprising an amino acid sequence selected from the group
consisting of SEQ ID NO:l-
48, b) a polypeptide comprising a naturally occurring amino acid sequence at
least 90% identical or at
least about 90% identical to an amino acid sequence selected from the group
consisting of SEQ )D
NO: l-48, c) a biologically active fragment of a polypeptide having an amino
acid sequence selected
from the group consisting of SEQ ~ NO:1-48, and d) an immunogenic fragment of
a polypeptide
having an amino acid sequence selected from the group consisting of SEQ ID
NO:1-48. Another
embodiment provides an isolated polypeptide comprising an amino acid sequence
of SEQ )D
NO:1-48.
Still another embodiment provides an isolated polynucleotide encoding a
polypeptide
selected from the group consisting of a) a polypeptide comprising an amino
acid sequence selected
from the group consisting of SEQ ~ N0:1-48, b) a polypeptide comprising a
naturally occurring
amino acid sequence at least 90% identical or at least about 90% identical to
an amino acid sequence
selected from the group consisting of SEQ >D NO:1-48, c) a biologically active
fragment of a
polypeptide having an amino acid sequence selected from the group consisting
of SEQ >D NO: l-48,
and d) an immunogenic fragment of a polypeptide having an amino acid sequence
selected from the
group consisting of SEQ ~ NO:1-48. In another embodiment, the polynucleotide
encodes a
polypeptide selected from the group consisting of SEQ >D NO: l-48. In an
alternative embodiment,
the polynucleotide is selected from the group consisting of SEQ ID N0:49-96.
Still another embodiment provides a recombinant polynucleotide comprising a
promoter
sequence operably linked to a polynucleotide encoding a polypeptide selected
from the group
4
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consisting of a) a polypeptide comprising an amino acid sequence selected from
the group consisting
of SEQ m NO:1-48, b) a polypeptide comprising a naturally occurring amino acid
sequence at least
90% identical or at least about 90% identical to an amino acid sequence
selected from the group
consisting of SEQ m NO:1-48, c) a biologically active fragment of a
polypeptide having an amino
acid sequence selected from the group consisting of SEQ ID NO: l-48, and d) an
ixnmunogenic
fragment of a polypeptide having an amino acid sequence selected from the
group consisting of SEQ
m NO:1-48. Another embodiment provides a cell transformed with the recombinant
polynucleotide.
Yet another embodiment provides a transgenic organism comprising the
recombinant polynucleotide.
Another embodiment provides a method for producing a polypeptide selected from
the group
consisting of a) a polypeptide comprising an amino acid sequence selected from
the group consisting
of SEQ )D NO:1-48, b) a polypeptide comprising a naturally occurring amino
acid sequence at least
90%o identical or at least about 90% identical to an amino acid sequence
selected from the group
consisting of SEQ ID NO:1-48, c) a biologically active fragment of a
polypeptide having an amino
acid sequence selected from the group consisting of SEQ m NO: l-48, and d) an
immunogenic
fragment of a polypeptide having an amino acid sequence selected from the
group consisting of SEQ
m NO: l-48. The method comprises a) culturing a cell under conditions suitable
for expression of the
polypeptide, wherein said cell is transformed with a recombinant
polynucleotide comprising a
promoter sequence operably linked to a polynucleotide encoding the
polypeptide, and b) recovering
the polypeptide so expressed.
Yet another embodiment provides an isolated antibody which specifically binds
to a
polypeptide selected from the group consisting of a) a polypeptide comprising
an amino acid
sequence selected from the group consisting of SEQ ID NO:1-48, b) a
polypeptide comprising a
naturally occurring amino acid sequence at least 90% identical or at least
about 90% identical to an
amino acid sequence selected from the group consisting of SEQ ID NO:1-48, c) a
biologically active
fragment of a polypeptide having an amino acid sequence selected from the
group consisting of SEQ
~ NO:1-48, and d) an immunogenic fragment of a polypeptide having an amino
acid sequence
selected from the group consisting of SEQ ID NO: l-48.
Still yet another embodiment provides an isolated polynucleotide selected from
the group
consisting of a) a polynucleotide comprising a polynucleotide sequence
selected from the group
consisting of SEQ )~ N0:49-96, b) a polynucleotide comprising a naturally
occurring polynucleotide
sequence at least 90% identical or at least about 90% identical to a
polynucleotide sequence selected
from the group consisting of SEQ )D N0:49-96, c) a polynucleotide
complementary to the
polynucleotide of a), d) a polynucleotide complementary to the polynucleotide
of b), and e) an RNA
equivalent of a)-d). In other embodiments, the polynucleotide can comprise at
least about 20, 30, 40,
60, 80, or 100 contiguous nucleotides.
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Yet another embodiment provides a method for detecting a target polynucleotide
in a sample,
said target polynucleotide being selected from the group consisting of a) a
polynucleotide comprising
a polynucleotide sequence selected from the group consisting of SEQ ID N0:49-
96, b) a
polynucleotide comprising a naturally occurring polynucleotide sequence at
least 90% identical or at
least about 90% identical to a polynucleotide sequence selected from the group
consisting of SEQ ID
N0:49-96, c) a polynucleotide complementary to the polynucleotide of a), d) a
polynucleotide
complementary to the polynucleotide of b), and e) an RNA equivalent of a)-d).
The method
comprises a) hybridizing the sample with a probe comprising at least 20
contiguous nucleotides
comprising a sequence complementary to said target polynucleotide in the
sample, and which probe
specifically hybridizes to said target polynucleotide, under conditions
whereby a hybridization
complex is formed between said probe and said target polynucleotide or
fragments thereof, and b)
detecting the presence or absence of said hybridization complex. In a related
embodiment, the
method can include detecting the amount of the hybridization complex. In still
other embodiments,
the probe can comprise at least about 20, 30, 40, 60, 80, or 100 contiguous
nucleotides.
Still yet another embodiment provides a method for detecting a target
polynucleotide in a
sample, said target polynucleotide being selected from the group consisting of
a) a polynucleotide
comprising a polynucleotide sequence selected from the group consisting of SEQ
ID N0:49-96, b) a
polynucleotide comprising a naturally occurring polynucleotide sequence at
least 90% identical or at
least about 90% identical to a polynucleotide sequence selected from the group
consisting of SEQ ID
N0:49-96, c) a polynucleotide complementary to the polynucleotide of a), d) a
polynucleotide
complementary to the polynucleotide of b), and e) an RNA equivalent of a)-d).
The method
comprises a) amplifying said target polynucleotide or fragment thereof using
polymerase chain
reaction amplification, and b) detecting the presence or absence of said
amplified target
polynucleotide or fragment thereof. In a related embodiment, the method can
include detecting the
amount of the amplified target polynucleotide or fragment thereof.
Another embodiment provides a composition comprising an effective amount of a
polypeptide selected from the group consisting of a) a polypeptide comprising
an amino acid
sequence selected from the group consisting of SEQ ID NO:1-48, b) a
polypeptide comprising a
naturally occurring amino acid sequence at least 90% identical or at least
about 90% identical to an
amino acid sequence selected from the group consisting of SEQ ID NO: l-48, c)
a biologically active
fragment of a polypeptide having an amino acid sequence selected from the
group consisting of SEQ
ID NO:1-48, and d) an immunogenic fragment of a polypeptide having an amino
acid sequence
selected from the group consisting of SEQ ID NO:1-48, and a pharmaceutically
acceptable excipient.
In one embodiment, the composition can comprise an amino acid sequence
selected from the group
consisting of SEQ ID NO:1-48. Other embodiments provide a method of treating a
disease or
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condition associated with decreased or abnormal expression of functional MDDT,
comprising
administering to a patient in need of such treatment the composition.
Yet another embodiment provides a method for screening a compound for
effectiveness as an
agonist of a polypeptide selected from the group consisting of a) a
polypeptide comprising an amino
acid sequence selected from the group consisting of SEQ ID NO:1-48, b) a
polypeptide comprising a
naturally occurring amino acid sequence at least 90% identical or at least
about 90% identical to an
amino acid sequence selected from the group consisting of SEQ ll~ N0:1-48, c)
a biologically active
fragment of a polypeptide having an amino acid sequence selected from the
group consisting of SEQ
)D NO:1-48, and d) an immunogenic fragment of a polypeptide having an amino
acid sequence
selected from the group consisting of SEQ ID N0:1-48. The method comprises a)
exposing a sample
comprising the polypeptide to a compound, and b) detecting agonist activity in
the sample. Another
embodiment provides a composition comprising an agonist compound identified by
the method and a
pharmaceutically acceptable excipient. Yet another embodiment provides a
method of treating a
disease or condition associated with decreased expression of functional MDDT,
comprising
administering to a patient in need of such treatment the composition.
Still yet another embodiment provides a method for screening a compound for
effectiveness
as an antagonist of a polypeptide selected from the group consisting of a) a
polypeptide comprising an
amino acid sequence selected from the group consisting of SEQ ll~ NO:1-48, b)
a polypeptide
comprising a naturally occurring amino acid sequence at least 90% identical or
at least about 90%
identical to an amino acid sequence selected from the group consisting of SEQ
ID NO:1-48, c) a
biologically active fragment of a polypeptide having an amino acid sequence
selected from the group
consisting of SEQ ID NO:1-48, and d) an immunogenic fragment of a polypeptide
having an amino
acid sequence selected from the group consisting of SEQ ID NO:l-48. The method
comprises a)
exposing a sample comprising the polypeptide to a compound, and b) detecting
antagonist activity in
the sample. Another embodiment provides a composition comprising an antagonist
compound
identified by the method and a pharmaceutically acceptable excipient. Yet
another embodiment
provides a method of treating a disease or condition associated with
overexpression of functional
MDDT, comprising administering to a patient in need of such treatment the
composition.
Another embodiment provides a method of screening for a compound that
specifically binds
to a polypeptide selected from the group consisting of a) a polypeptide
comprising an amino acid
sequence selected from the group consisting of SEQ ll~ NO: l-48, b) a
polypeptide comprising a
naturally occurring amino acid sequence at least 90% identical or at least
about 90% identical to an
amino acid sequence selected from the group consisting of SEQ m NO:1-48, c) a
biologically active
fragment of a polypeptide having an amino acid sequence selected from the
group consisting of SEQ
)D NO:1-48, and d) an immunogenic fragment of a polypeptide having an amino
acid sequence
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selected from the group consisting of SEQ. DJ NO:1-48. The method comprises a)
combining the
polypeptide with at least one test compound under suitable conditions, and b)
detecting binding of the
polypeptide to the test compound, thereby identifying a compound that
specifically binds to the
polypeptide.
Yet another embodiment provides a method of screening for a compound that
modulates the
activity of a polypeptide selected from the group consisting of a) a
polypeptide comprising an amino
acid sequence selected from the group consisting of SEQ m NO:1-48, b) a
polypeptide comprising a
naturally occurring amino acid sequence at least 90% identical or at least
about 90% identical to an
amino acid sequence selected from the group consisting of SEQ m NO:1-48, c) a
biologically active
fragment of a polypeptide having an amino acid sequence selected from the
group consisting of SEQ
m N0:1-48, and d) an immunogenic fragment of a polypeptide having an amino
acid sequence
selected from the group consisting of SEQ ID NO: l-48. The method comprises a)
combining the
polypeptide with at least one test compound under conditions permissive for
the activity of the
polypeptide, b) assessing the activity of the polypeptide in the presence of
the test compound, and c)
comparing the activity of the polypeptide in the presence of the test compound
with the activity of the
polypeptide in the absence of the test compound, wherein a change in the
activity of the polypeptide
in the presence of the test compound is indicative of a compound that
modulates the activity of the
polypeptide.
Still yet another embodiment provides a method for screening a compound for
effectiveness
in altering expression of a target polynucleotide, wherein said target
polynucleotide comprises a
polynucleotide sequence selected from the group consisting of SEQ )D N0:49-96,
the method
comprising a) exposing a sample comprising the target polynucleotide to a
compound, b) detecting
altered expression of the target polynucleotide, and c) comparing the
expression of the target
polynucleotide in the presence of varying amounts of the compound and in the
absence of the
compound.
Another embodiment provides a method for assessing toxicity of a test
compound, said
method comprising a) treating a biological sample containing nucleic acids
with the test compound;
b) hybridizing the nucleic acids of the treated biological sample with a probe
comprising at least 20
contiguous nucleotides of a polynucleotide selected from the group consisting
of i) a polynucleotide
comprising a polynucleotide sequence selected from the group consisting of SEQ
)D N0:49-96, ii) a
polynucleotide comprising a naturally occurnng polynucleotide sequence at
least 90% identical or at
least about 90% identical to a polynucleotide sequence selected from the group
consisting of SEQ m
N0:49-96, iii) a polynucleotide having a sequence complementary to i), iv) a
polynucleotide
complementary to the polynucleotide of ii), and v) an RNA equivalent of i)-
iv). Hybridization occurs
under conditions whereby a specific hybridization complex is formed between
said probe and a target
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polynucleotide in the biological sample, said target polynucleotide selected
from the group consisting
of i) a polynucleotide comprising a polynucleotide sequence selected from the
group consisting of
SEQ )I7 N0:49-96, ii) a polynucleotide comprising a naturally occurring
polynucleotide sequence at
least 90% identical or at least about 90% identical to a polynucleotide
sequence selected from the
group consisting of SEQ ID N0:49-96, iii) a polynucleotide complementary to
the polynucleotide of
i), iv) a polynucleotide complementary to the polynucleotide of ii), and v) an
RNA equivalent of i)-
iv). Alternatively, the target polynucleotide can comprise a fragment of a
polynucleotide selected
from the group consisting of i)-v) above; c) quantifying the amount of
hybridization complex; and d)
comparing the amount of hybridization complex in the treated biological sample
with the amount of
hybridization complex in an untreated biological sample, wherein a difference
in the amount of
hybridization complex in the treated biological sample is indicative of
toxicity of the test compound.
BRIEF DESCRIPTION OF THE TABLES
Table 1 summarizes the nomenclature for full length polynucleotide and
polypeptide
embodiments of the invention.
Table 2 shows the GenBank identification number and annotation of the nearest
GenBank
homolog, and the PROTEOME database identification numbers and annotations of
PROTEOME
database homologs, for polypeptide embodiments of the invention. The
probability scores for the
matches between each polypeptide and its homolog(s) are also shown.
\~ 20 Table 3 shows structural features of polypeptide embodiments, including
predicted motifs
and domains, along with the methods, algorithms, and searchable databases used
for analysis of the
polypeptides.
Table 4 lists the cDNA and/or genomic DNA fragments which were used to
assemble
polynucleotide embodiments, along with selected fragments of the
polynucleotides.
Table 5 shows representative cDNA libraries for polynucleotide embodiments.
Table 6 provides an appendix which describes the tissues and vectors used for
construction of
the cDNA libraries shown in Table 5
Table 7 shows the tools, programs, and algorithms used to analyze
polynucleotides and
polypeptides, along with applicable descriptions, references, and threshold
parameters.
DESCRIPTION OF THE INVENTION
Before the present proteins, nucleic acids, and methods are described, it is
understood that
embodiments of the invention are not limited to the particular machines,
instruments, materials, and
methods described, as these may vary. It is also to be understood that the
terminology used herein is
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for the purpose of describing particular embodiments only, and is not intended
to limit the scope of
the invention.
As used herein and in the appended claims, the singular forms "a," "an," and
"the" include
plural reference unless the context clearly dictates otherwise. Thus, for
example, a reference to "a
host cell" includes a plurality of such host cells, and a reference to "an
antibody" is a reference to one
or more antibodies and equivalents thereof known to those skilled in the art,
and so forth.
Unless defined otherwise, all technical and scientific terms used herein have
the same
meanings as commonly understood by one 'of ordinary skill in the art to which
this invention belongs.
Although any machines, materials, and methods similar or equivalent to those
described herein can be
used to practice or test the present invention, the preferred machines,
materials and methods are now
described. All publications mentioned herein are cited for the purpose of
describing and disclosing
the cell lines, protocols, reagents and vectors which are reported in the
publications and which might
be used in connection with various embodiments of the invention. Nothing
herein is to be construed
as an admission that the invention is not entitled to antedate such disclosure
by virtue of prior
invention.
DEFINITIONS
"MDDT" refers to the amino acid sequences of substantially purified MDDT
obtained from
any species, particularly a mammalian species, including bovine, ovine,
porcine, murine, equine, and
human, and from any source, whether natural, synthetic, semi-synthetic, or
recombinant.
The term "agonist" refers to a molecule which intensifies or mimics the
biological activity of
MDDT. Agonists may include proteins, nucleic acids, carbohydrates, small
molecules, or any other
compound or composition which modulates the activity of MDDT either by
directly interacting with
MDDT or by acting on components of the biological pathway in which MDDT
participates.
An "allelic variant" is an alternative form of the gene encoding MDDT. Allelic
variants may
result from at least one mutation in the nucleic acid sequence and may result
in altered mRNAs or in
polypeptides whose structure or function may or may not be altered. A gene may
have none, one, or
many allelic variants of its naturally occurring form. Common mutational
changes which give rise to
allelic variants are generally ascribed to natural deletions, additions, or
substitutions of nucleotides.
Each of these types of changes may occur alone, or in combination with the
others, one or more times
in a given sequence.
"Altered" nucleic acid sequences encoding MDDT include those sequences with
deletions,
insertions, or substitutions of different nucleotides, resulting in a
polypeptide the same as MDDT or a
polypeptide with at least one functional characteristic of MDDT. Included
within this definition are
polymorphisms which may or may not be readily detectable using a particular
oligonucleotide probe
of the polynucleotide encoding MDDT, and improper or unexpected hybridization
to allelic variants,
1o
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with a locus other than the normal chromosomal locus for the polynucleotide
encoding MDDT. The
encoded protein may also be "altered," and may contain deletions, insertions,
or substitutions of
amino acid residues which produce a silent change and result in a functionally
equivalent MDDT.
Deliberate amino acid substitutions may be made on the basis of one or more
similarities in polarity,
charge, solubility, hydrophobicity, hydrophilicity, and/or the amphipathic
nature of the residues, as
long as the biological or immunological activity of MDDT is retained. For
example, negatively
charged amino acids may include aspartic acid and glutamic acid, and
positively charged amino acids
may include lysine and arginine. Amino acids with uncharged polar side chains
having similar
hydrophilicity values may include: asparagine and glutamine; and serine and
threonine. Amino acids
with uncharged side chains having similar hydrophilicity values may include:
leucine, isoleucine, and
valine; glycine and alanine; and phenylalanine and tyrosine.
The terms "amino acid" and "amino acid sequence" can refer to an oligopeptide,
a peptide, a
polypeptide, or a protein sequence, or a fragment of any of these, and to
naturally occurring or
synthetic molecules. Where "amino acid sequence" is recited to refer to a
sequence of a naturally
occurring protein molecule, "amino acid sequence" and like terms are not meant
to limit the amino
acid sequence to the complete native amino acid sequence associated with the
recited protein
molecule.
"Amplification" relates to the production of additional copies of a nucleic
acid.
Amplification may be carried out using polymerase chain reaction (PCR)
technologies or other
nucleic acid amplification technologies well known in the art.
The term "antagonist" refers to a molecule which inhibits or attenuates the
biological activity
of MDDT. Antagonists may include proteins such as antibodies, anticalins,
nucleic acids,
carbohydrates, small molecules, or any other compound or composition which
modulates the activity
of MDDT either by directly interacting with MDDT or by acting on components of
the biological
pathway in which MDDT participates.
The term "antibody" refers to intact immunoglobulin molecules as well as to
fragments
thereof, such as Fab, F(ab')~, and Fv fragments, which are capable of binding
an epitopic determinant.
Antibodies that bind MDDT polypeptides can be prepared using intact
polypeptides or using
fragments containing small peptides of interest as the immunizing antigen. The
polypeptide or
oligopeptide used to immunize an animal (e.g., a mouse, a rat, or a rabbit)
can be derived from the
translation of RNA, or synthesized chemically, and can be conjugated to a
carrier protein if desired.
Commonly used carriers that are chemically coupled to peptides include bovine
serum albumin,
thyroglobulin, and keyhole limpet hemocyanin (KLH). The coupled peptide is
then used to immunize
the animal.
The term "antigenic determinant" refers to that region of a molecule (i.e., an
epitope) that
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makes contact with a particular antibody. When a protein or a fragment of a
protein is used to
immunize a host animal, numerous regions of the protein may induce the
production of antibodies
which bind specifically to antigenic determinants (particular regions or three-
dimensional structures
on the protein). An antigenic determinant may compete with the intact antigen
(i.e., the immunogen
used to elicit the immune response) for binding to an antibody.
The term "aptamer" refers to a nucleic acid or oligonucleotide molecule that
binds to a
specific molecular target. Aptamers are derived from an in vitro evolutionary
process (e.g., SELEX
(Systematic Evolution of Ligands by EXponential Enrichment), described in U.S.
Patent No.
5,270,163), which selects for target-specific aptamer sequences from large
combinatorial libraries.
Aptamer compositions may be double-stranded or single-stranded, and may
include
deoxyribonucleotides, ribonucleotides, nucleotide derivatives, or other
nucleotide-like molecules.
The nucleotide components of an aptamer may have modified sugar groups (e.g.,
the 2'-OH group of a
ribonucleotide may be replaced by 2'-F or 2'-NHZ), which may improve a desired
property, e.g.,
resistance to nucleases or longer lifetime in blood. Aptamers may be
conjugated to other molecules,
e.g., a high molecular weight carrier to slow clearance of the aptamer from
the circulatory system.
Aptamers may be specifically cross-linked to their cognate ligands, e.g., by
photo-activation of a
cross-linker (Brody, E.N. and L. Gold (2000) J. Biotechnol. 74:5-13).
The term "intramer" refers to an aptamer which is expressed irz vivo. For
example, a vaccinia
virus-based RNA expression system has been used to express specific RNA
aptamers at high levels in
the cytoplasm of leukocytes (Blind, M. et al. (1999) Proc. Natl. Acad. Sci.
USA 96:3606-3610).
The term "spiegelmer" refers to an aptamer which includes L-DNA, L-RNA, or
other left-
handed nucleotide derivatives or nucleotide-like molecules. Aptamers
containing left-handed
nucleotides are resistant to degradation by naturally occurring enzymes, which
normally act on
substrates containing right-handed nucleotides.
The term "antisense" refers to any composition capable of base-pairing with
the "sense"
(coding) strand of a polynucleotide having a specific nucleic acid sequence.
Antisense compositions
may include DNA; RNA; peptide nucleic acid (PNA); oligonucleotides having
modified backbone
linkages such as phosphorothioates, methylphosphonates, or benzylphosphonates;
oligonucleotides
having modified sugar groups such as 2'-methoxyethyl sugars or 2'-
methoxyethoxy sugars; or
oligonucleotides having modified bases such as 5-methyl cytosine, 2'-
deoxyuracil, or 7-deaza-2'-
deoxyguanosine. Antisense molecules may be produced by any method including
chemical synthesis
or transcription. Once introduced into a cell, the complementary antisense
molecule base-pairs with a
naturally occurring nucleic acid sequence produced by the cell to form
duplexes which block either
transcription or translation. The designation "negative" or "minus" can refer
to the antisense strand,
and the designation "positive" or "plus" can refer to the sense strand of a
reference DNA molecule.
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The term "biologically active" refers to a protein having structural,
regulatory, or biochemical
functions of a naturally occurring molecule. Likewise, "immunologically
active" or "immunogenic"
refers to the capability of the natural, recombinant, or synthetic MDDT, or of
any oligopeptide
thereof, to induce a specific immune response in appropriate animals or cells
and to bind with specific
antibodies.
"Complementary" describes the relationship between two single-stranded nucleic
acid
sequences that anneal by base-pairing. For example, 5'-AGT-3' pairs with its
complement,
3'-TCA-5'.
A "composition comprising a given polynucleotide" and a "composition
comprising a given
polypeptide" can refer to any composition containing the given polynucleotide
or polypeptide. The
composition may comprise a dry formulation or an aqueous solution.
Compositions comprising
polynucleotides encoding MDDT or fragments of MDDT may be employed as
hybridization probes.
The probes may be stored in freeze-dried form and may be associated with a
stabilizing agent such as
a carbohydrate. In hybridizations, the probe may be deployed in an aqueous
solution containing salts
(e.g., NaCl), detergents (e.g., sodium dodecyl sulfate; SDS), and other
components (e.g., Denhardt's
solution, dry milk, salmon sperm DNA, etc.).
"Consensus sequence" refers to a nucleic acid sequence which has been
subjected to repeated
DNA sequence analysis to resolve uncalled bases, extended using the XL-PCR kit
(Applied
Biosystems, Foster City CA) in the 5' and/or the 3' direction, and
resequenced, or which has been
assembled from one or more overlapping cDNA, EST, or genomic DNA fragments
using a computer
program for fragment assembly, such as the GELVIEW fragment assembly system
(Accelrys,
Burlington MA) or Phrap (University of Washington, Seattle WA). Some sequences
have been both
extended and assembled to produce the consensus sequence.
"Conservative amino acid substitutions" are those substitutions that are
predicted to least
interfere with the properties of the original protein, i.e., the structure and
especially the function of
the protein is conserved and not significantly changed by such substitutions.
The table below shows
amino acids which may be substituted for an original amino acid in a protein
and which are regarded
as conservative amino acid substitutions.
Original Residue Conservative Substitution
Ala Gly, Ser
Arg His, Lys
Asn Asp, Gln, His
Asp Asn, Glu
Cys Ala, Ser
Gln Asn, Glu, His
Glu Asp, Gln, His
Gly Ala
His Asn, Arg, Gln, Glu
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Ile Leu, Val
Leu Ile, Val
Lys Arg, Gln, Glu
Met Leu, Ile
Phe His, Met, Leu, Trp, Tyr
Ser Cys, Thr
Thr Ser, Val
Trp Phe, Tyr
Tyr His, Phe, Trp
Val Ile, Leu, Thr
Conservative amino acid substitutions generally maintain (a) the structure of
the polypeptide
backbone in the area of the substitution, for example, as a beta sheet or
alpha helical conformation,
(b) the charge or hydrophobicity of the molecule at the site of the
substitution, and/or (c) the bulk of
the side chain.
A "deletion" refers to a change in the amino acid or nucleotide sequence that
results in the
absence of one or more amino acid residues or nucleotides.
The term "derivative" refers to a chemically modified polynucleotide or
polypeptide.
Chemical modifications of a polynucleotide can include, for example,
replacement of hydrogen by an
alkyl, acyl, hydroxyl, or amino group. A derivative polynucleotide encodes a
polypeptide which
retains at least one biological or immunological function of the natural
molecule. A derivative
polypeptide is one modified by glycosylation, pegylation, or any similar
process that retains at least
one biological or immunological function of the polypeptide from which it was
derived.
A "detectable label" refers to a reporter molecule or enzyme that is capable
of generating a
measurable signal and is covalently or noncovalently joined to a
polynucleotide or polypeptide.
"Differential expression" refers to increased or upregulated; or decreased,
downregulated, or
absent gene or protein expression, determined by comparing at least two
different samples. Such
comparisons may be carned out between, for example, a treated and an untreated
sample, or a
diseased and a normal sample.
"Exon shuffling" refers to the recombination of different coding regions
(exons). Since an
exon may represent a structural or functional domain of the encoded protein,
new proteins may be
assembled through the novel reassortment of stable substructures, thus
allowing acceleration of the
evolution of new protein functions.
A "fragment" is a unique portion of MDDT or a polynucleotide encoding MDDT
which can
be identical in sequence to, but shorter in length than, the parent sequence.
A fragment may comprise
up to the entire length of the defined sequence, minus one nucleotide/amino
acid residue. For
example, a fragment may comprise from about 5 to about 1000~contiguous
nucleotides or amino acid
residues. A fragment used as a probe, primer, antigen, therapeutic molecule,
or for other purposes,
may be at least 5, 10, 15, 16, 20, 25, 30, 40, 50, 60, 75, 100, 150, 250 or at
least 500 contiguous
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nucleotides or amino acid residues in length. Fragments may be preferentially
selected from certain
regions of a molecule. For example, a polypeptide fragment may comprise a
certain length of
contiguous amino acids selected from the first 250 or 500 amino acids (or
first 25% or 50%) of a
polypeptide as shown in a certain defined sequence. Clearly these lengths are
exemplary, and any
length that is supported by the specification, including the Sequence Listing,
tables, and figures, may
be encompassed by the present embodiments.
A fragment of SEQ ID N0:49-96 can comprise a region of unique polynucleotide
sequence
that specifically identifies SEQ ID N0:49-96, for example, as distinct from
any other sequence in the
genome from which the fragment was obtained. A fragment of SEQ )D N0:49-96 can
be employed
in one or more embodiments of methods of the invention, for example, in
hybridization and
amplification technologies and in analogous methods that distinguish SEQ ID
N0:49-96 from related
polynucleotides. The precise length of a fragment of SEQ )D N0:49-96 and the
region of SEQ )D
N0:49-96 to which the fragment corresponds are routinely determinable,by one
of ordinary skill in
the art based on the intended purpose for the fragment.
A fragment of SEQ )D NO:1-48 is encoded by a fragment of SEQ ID N0:49-96. A
fragment
of SEQ ~ NO: l-48 can comprise a region of unique amino acid sequence that
specifically identifies
SEQ ID N0:1-48. For example, a fragment of SEQ ID NO:1-48 can be used as an
immunogenic
peptide for the development of antibodies that specifically recognize SEQ ID
NO:1-48. The precise
length of a fragment of SEQ ~ NO:1-48 and the region of SEQ ID NO: l-48 to
which the fragment
corresponds can be determined based on the intended purpose for the fragment
using one or more
analytical methods described herein or otherwise known in the art.
A "full length" polynucleotide is one containing at least a translation
initiation codon (e.g.,
methionine) followed by an open reading frame and a translation termination
codon. A "full length"
polynucleotide sequence encodes a "full length" polypeptide sequence.
"Homology" refers to sequence similarity or, alternatively, sequence identity,
between two or
more polynucleotide sequences or two or more polypeptide sequences.
The terms "percent identity" and "% identity," as applied to polynucleotide
sequences, refer
to the percentage of identical residue matches between at least two
polynucleotide sequences aligned
using a standardized algorithm. Such an algorithm may insert, in a
standardized and reproducible
way, gaps in the sequences being compared in order to optimize alignment
between two sequences,
and therefore achieve a more meaningful comparison of the two sequences.
Percent identity between polynucleotide sequences may be determined using one
or more
computer algorithms or programs known in the art or described herein. For
example, percent identity
can be determined using the default parameters of the CLUSTAL V algorithm as
incorporated into
the MEGALIGN version 3.12e sequence alignment program. This program is part of
the
CA 02460480 2004-03-15
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LASERGENE software package, a suite of molecular biological analysis programs
(DNASTAR,
Madison WI). CLUSTAL V is described in Higgins, D.G. and P.M. Sharp (1989;
CABIOS 5:151-
153) and in Higgins, D.G. et al. (1992; CABIOS 8:189-191). For pairwise
alignments of
polynucleotide sequences, the default parameters are set as follows: Ktuple=2,
gap penalty=5,
window=4, and "diagonals saved"=4. The "weighted" residue weight table is
selected as the default.
Alternatively, a suite of commonly used and freely available sequence
comparison algorithms
which can be used is provided by the National Center for Biotechnology
Information (NCBI) Basic
Local Alignment Search Tool (BLAST) (Altschul, S.F. et al. (1990) J. Mol.
Biol. 215:403-410),
which is available from several sources, including the NCBI, Bethesda, MD, and
on the Internet at
http://www.ncbi.nlm.nih.gov/BLAST/. The BLAST software suite includes various
sequence
analysis programs including "blastn," that is used to align a known
polynucleotide sequence with
other polynucleotide sequences from a variety of databases. Also available is
a tool called "BLAST 2
Sequences" that is used for direct pairwise comparison of two nucleotide
sequences. "BLAST 2
Sequences" can be accessed and used interactively at
http://www.ncbi.nlm.nih.gov/gorf/bl2.html.
The "BLAST 2 Sequences" tool can be used for both blastn and blastp (discussed
below). BLAST
programs are commonly used with gap and other parameters set to default
settings. For example, to
compare two nucleotide sequences, one may use blastn with the "BLAST 2
Sequences" tool Version
2Ø12 (April-21-2000) set at default parameters. Such default parameters may
be, for example:
Matrix: BLOSUM62
Reward for- match: 1
Penalty for mismatch: -2
Opezz Gap: 5 and Extezzsiozz Gap: 2 penalties
Gap x drop-off.' S0
Expect: 10
Word Size: 11
Filter: on
Percent identity may be measured over the length of an entire defined
sequence, for example,
as defined by a particular SEQ ID number, or may be measured over a shorter
length, for example,
over the length of a fragment taken from a larger, defined sequence, for
instance, a fragment of at
least 20, at least 30, at least 40, at least 50, at least 70, at least 100, or
at least 200 contiguous
nucleotides. Such lengths are exemplary only, and it is understood that any
fragment length
supported by the sequences shown herein, in the tables, figures, or Sequence
Listing, may be used to
describe a length over which percentage identity may be measured.
Nucleic acid sequences that do not show a high degree of identity may
nevertheless encode
similar amino acid sequences due to the degeneracy of the genetic code. It is
understood that changes
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in a nucleic acid sequence can be made using this degeneracy to produce
multiple nucleic acid
sequences that all encode substantially the same protein.
The phrases "percent identity" and "% identity," as applied to polypeptide
sequences, refer to
the percentage of identical residue matches between at least two polypeptide
sequences aligned using
a standardized algorithm. Methods of polypeptide sequence alignment are well-
known. Some
alignment methods take into account conservative amino acid substitutions.
Such conservative
substitutions, explained in more detail above, generally preserve the charge
and hydrophobicity at the
site of substitution, thus preserving the structure (and therefore function)
of the polypeptide. The
phrases "percent similarity" and "% similarity," as applied to polypeptide
sequences, refer to the
percentage of residue matches, including identical residue matches and
conservative substitutions,
between at least two polypeptide sequences aligned using a standardized
algorithm. In contrast,
conservative substitutions are not included in the calculation of percent
identity between polypeptide
sequences.
Percent identity between polypeptide sequences may be determined using the
default
parameters of the CLUSTAL V algorithm as incorporated into the MEGALIGN
version 3.12e
sequence alignment program (described and referenced above). For pairwise
alignments of
polypeptide sequences using CLUSTAL V, the default parameters are set as
follows: I~tuple=1, gap
penalty=3, window=5, and "diagonals saved"=5. The PAM250 matrix is selected as
the default
residue weight table.
Alternatively the NCBI BLAST software suite may be used. For example, for a
pairwise
comparison of two polypeptide sequences, one may use the "BLAST 2 Sequences"
tool Version
2Ø12 (April-21-2000) with blastp set at default parameters. Such default
parameters may be, for
example:
Matrix: BLOSUM62
Open Gap: Il and Exterzsio~a Gap: I penalties
Gap x drop-off.' S0
Expect: 10
Word Size: 3
Filter: oh
Percent identity may be measured over the length of an entire defined
polypeptide sequence,
for example, as defined by a particular SEQ ID number, or may be measured over
a shorter length, for
example, over the length of a fragment taken from a larger, defined
polypeptide sequence, for
instance, a fragment of at least 15, at least 20, at least 30, at least 40, at
least 50, at least 70 dr at least
150 contiguous residues. Such lengths are exemplary only, and it is understood
that any fragment
length supported by the sequences shown herein, in the tables, figures or
Sequence Listing, may be
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used to describe a length over which percentage identity may be measured.
"Human artificial chromosomes" (HACs) are linear microchromosomes which may
contain
DNA sequences of about 6 kb to 10 Mb in size and which contain all of the
elements required for
chromosome replication, segregation and maintenance.
The term "humanized antibody" refers to an antibody molecule in which the
amino acid
sequence in the non-antigen binding regions has been altered so that the
antibody more closely
resembles a human antibody, and still retains its original binding ability.
"Hybridization" refers to the process by which a polynucleotide strand anneals
with a
complementary strand through base pairing under defined hybridization
conditions. Specific
hybridization is an indication that two nucleic acid sequences share a high
degree of
complementarity. Specific hybridization complexes form under permissive
annealing conditions and
remain hybridized after the "washing" step(s). The washing steps) is
particularly important in
determining the stringency of the hybridization process, with more stringent
conditions allowing less
non-specific binding, i.e., binding between pairs of nucleic acid strands that
are not perfectly
matched. Permissive conditions for annealing of nucleic acid sequences are
routinely determinable
by one of ordinary skill in the art and may be consistent among hybridization
experiments, whereas
wash conditions may be varied among experiments to achieve the desired
stringency, and therefore
hybridization specificity. Permissive annealing conditions occur, for example,
at 68°C in the
presence of about 6 x SSC, about 1% (w/v) SDS, and about 100 ~.g/ml sheared,
denatured salmon
sperm DNA.
Generally, stringency of hybridization is expressed, in part, with reference
to the temperature
under which the wash step is carried out. Such wash temperatures are typically
selected to be about
5°C to 20°C lower than the thermal melting point (T",) for the
specific sequence at a defined ionic
strength and pH. The T,m is the temperature (under defined ionic strength and
pH) at which 50% of
the target sequence hybridizes to a perfectly matched probe. An equation for
calculating Tm and
conditions for nucleic acid hybridization are well known and can be found in
Sambrook, J. and D.W.
Russell (2001; Molecular Cloning: A Laboratory Manual, 3rd ed., vol. 1-3, Cold
Spring Harbor Press,
Cold Spring Harbor NY, ch. 9).
High stringency conditions for hybridization between polynucleotides of the
present
invention include wash conditions of 68°C in the presence of about 0.2
x SSC and about 0.1% SDS,
for 1 hour. Alternatively, temperatures of about 65°C, 60°C,
55°C, or 42°C may be used. SSC
concentration may be varied from about 0.1 to 2 x SSC, with SDS being present
at about 0.1%.
Typically, blocking reagents are used to block non-specific hybridization.
Such blocking reagents
include, for instance, sheared and denatured salmon sperm DNA at about 100-200
~.g/ml. Organic
solvent, such as formamide at a concentration of about 35-50% v/v, may also be
used under particular
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circumstances, such as for RNA:DNA hybridizations. Useful variations on these
wash conditions
will be readily apparent to those of ordinary skill in the art. Hybridization,
particularly under high
stringency conditions, may be suggestive of evolutionary similarity between
the nucleotides. Such
similarity is strongly indicative of a similar role for the nucleotides and
their encoded polypeptides.
The term "hybridization complex" refers to a complex formed between two
nucleic acids by
virtue of the formation of hydrogen bonds between complementary bases. A
hybridization complex
may be formed in solution (e.g., Cot or Rot analysis) or formed between one
nucleic acid present in
solution and another nucleic acid immobilized on a solid support (e.g., paper,
membranes, filters,
chips, pins or glass slides, or any other appropriate substrate to which cells
or their nucleic acids have
been fixed).
The words "insertion" and "addition" refer to changes in an amino acid or
polynucleotide
sequence resulting in the addition of one or more amino acid residues or
nucleotides, respectively.
"Immune response" can refer to conditions associated with inflammation,
trauma, immune
disorders, or infectious or genetic disease, etc. These conditions can be
characterized by expression
of various factors, e.g., cytokines, chemokines, and other signaling
molecules, which may affect
cellular and systemic defense systems.
An "immunogenic fragment" is a polypeptide or oligopeptide fragment of MDDT
which is
capable of eliciting an immune response when introduced into a living
organism, for example, a
marrunal. The term "immunogenic fragment" also includes any polypeptide or
oligopeptide fragment
of MDDT which is useful in any of the antibody production methods disclosed
herein or known in the
art.
The term "microarray" refers to an arrangement of a plurality of
polynucleotides,
polypeptides, antibodies, or other chemical compounds on a substrate.
The terms "element" and "array element" refer to a polynucleotide,
polypeptide, antibody, or
other chemical compound having a unique and defined position on a microarray.
The term "modulate" refers to a change in the activity of MDDT. For example,
modulation
may cause an increase or a decrease in protein activity, binding
characteristics, or any other
biological, functional, or immunological properties of MDDT.
The phrases "nucleic acid" and "nucleic acid sequence" refer to a nucleotide,
oligonucleotide,
polynucleotide, or any fragment thereof. These phrases also refer to DNA or
RNA of genomic or
synthetic origin which may be single-stranded or double-stranded and may
represent the sense or the
antisense strand, to peptide nucleic acid (PNA), or to any DNA-like or RNA-
like material.
"Operably linked" refers to the situation in which a first nucleic acid
sequence is placed in a
functional relationship with a second nucleic acid sequence. For instance, a
promoter is operably
linked to a coding sequence if the promoter affects the transcription or
expression of the coding
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sequence. Operably linked DNA sequences may be in close proximity or
contiguous and, where
necessary to join two protein coding regions, in the same reading frame.
"Peptide nucleic acid" (PNA) refers to an antisense molecule or anti-gene
agent which
comprises an oligonucleotide of at least about 5 nucleotides in length linked
to a peptide backbone of
amino acid residues ending in lysine. The terminal lysine confers solubility
to the composition.
PNAs preferentially bind complementary single stranded DNA or RNA and stop
transcript
elongation, and may be pegylated to extend their lifespan in the cell.
"Post-translational modification" of an MDDT may involve lipidation,
glycosylation,
phosphorylation, acetylation, racemization, proteolytic cleavage, and other
modifications known in
the art. These processes may occur synthetically or biochemically. Biochemical
modifications will
vary by cell type depending on the enzymatic milieu of MDDT.
"Probe" refers to nucleic acids encoding MDDT, their complements, or fragments
thereof,
which are used to detect identical, allelic or related nucleic acids. Probes
are isolated
oligonucleotides or polynucleotides attached to a detectable label or reporter
molecule. Typical
labels include radioactive isotopes, ligands, chemiluminescent agents, and
enzymes. "Primers" are
short nucleic acids, usually DNA oligonucleotides, which may be annealed to a
target polynucleotide
by complementary base-pairing. The primer may then be extended along the
target DNA strand by a
DNA polymerase enzyme. Primer pairs can be used for amplification (and
identification) of a nucleic
acid, e.g., by the polymerase chain reaction (PCR).
Probes and primers as used in the present invention typically comprise at
least 15 contiguous
nucleotides of a known sequence. In order to enhance specificity, longer
probes and primers may also
be employed, such as probes and primers that comprise at least 20, 25, 30, 40,
50, 60, 70, 80, 90, 100,
or at least 150 consecutive nucleotides of the disclosed nucleic acid
sequences. Probes and primers
may be considerably longer than these examples, and it is understood that any
length supported by the
specification, including the tables, figures, and Sequence Listing, may be
used.
Methods for preparing and using probes and primers are described in, for
example,
Sambrook, J. and D.W. Russell (2001; Molecular Cloning: A Laboratory Manual,
3rd ed., vol. 1-3,
Cold Spring Harbor Press, Cold Spring Harbor NY), Ausubel, F.M. et al. (1999;
Short Protocols in
Molecular Biolo~y, 4''' ed., John Wiley & Sons, New York NY), and Innis, M. et
al. (1990; PCR
Protocols, A Guide to Methods and Applications, Academic Press, San Diego CA).
PCR primer pairs
can be derived from a known sequence, for example, by using computer programs
intended for that
purpose such as Primer (Version 0.5, 1991, Whitehead Institute for Biomedical
Research, Cambridge
MA).
Oligonucleotides for use as primers are selected using software known in the
art for such
purpose. For example, OLIGO 4.06 software is useful for the selection of PCR
primer pairs of up to
CA 02460480 2004-03-15
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100 nucleotides each, and for the analysis of oligonucleotides and larger
polynucleotides of up to
5,000 nucleotides from an input polynucleotide sequence of up to 32 kilobases.
Similar primer
selection programs have incorporated additional features for expanded
capabilities. For example, the
PrimOU primer selection program (available to the public from the Genome
Center at University of
Texas South West Medical Center, Dallas TX) is capable of choosing specific
primers from
megabase sequences and is thus useful for designing primers on a genome-wide
scope. The Primer3
primer selection program (available to the public from the Whitehead
Institute/MIT Center for
Genome Research, Cambridge MA) allows the user to input a "mispriming
library," in which
sequences to avoid as primer binding sites are user-specified. Primer3 is
useful, in particular, for the
selection of oligonucleotides for microarrays. (The source code for the latter
two primer selection
programs may also be obtained from their respective sources and modified to
meet the user's specific
needs.) The PrimeGen program (available to the public from the UK Human Genome
Mapping
Project Resource Centre, Cambridge UI~) designs primers based on multiple
sequence alignments,
thereby allowing selection of primers that hybridize to either the most
conserved or least conserved
regions of aligned nucleic acid sequences. Hence, this program is useful for
identification of both
unique and conserved oligonucleotides and polynucleotide fragments. The
oligonucleotides and
polynucleotide fragments identified by any of the above selection methods are
useful in hybridization
technologies, for example, as PCR or sequencing primers, microarray elements,
or specific probes to
identify fully or partially complementary polynucleotides in a sample of
nucleic acids. Methods of
oligonucleotide selection are not limited to those described above.
A "recombinant nucleic acid" is a nucleic acid that is not naturally occurring
or has a
sequence that is made by an artificial combination of two or more otherwise
separated segments of
sequence. This artificial combination is often accomplished by chemical
synthesis or, more
commonly, by the artificial manipulation of isolated segments of nucleic
acids, e.g., by genetic
engineering techniques such as those described in Sambrook and Russell
(supra). The term
recombinant includes nucleic acids that have been altered solely by addition,
substitution, or deletion
of a portion of the nucleic acid. Frequently, a recombinant nucleic acid may
include a nucleic acid
sequence operably linked to a promoter sequence. Such a recombinant nucleic
acid may be part of a
vector that is used, for example, to transform a cell.
Alternatively, such recombinant nucleic acids may be part of a viral vector,
e.g., based on a
vaccinia virus, that could be use to vaccinate a mammal wherein the
recombinant nucleic acid is
expressed, inducing a protective immunological response in the mammal.
A "regulatory element" refers to a nucleic acid sequence usually derived from
untranslated
regions of a gene and includes enhancers, promoters, introns, and 5' and 3'
untranslated regions
(UTRs). Regulatory elements interact with host or viral proteins which control
transcription,
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translation, or RNA stability.
"Reporter molecules" are chemical or biochemical moieties used for labeling a
nucleic acid,
amino acid, or antibody. Reporter molecules include radionuclides; enzymes;
fluorescent,
chemiluminescent, or chromogenic agents; substrates; cofactors; inhibitors;
magnetic particles; and
other moieties known in the art.
An "RNA equivalent," in reference to a DNA molecule, is composed of the same
linear
sequence of nucleotides as the reference DNA molecule with the exception that
all occurrences of the
nitrogenous base thymine are replaced with uracil, and the sugar backbone is
composed of ribose
instead of deoxyribose.
The term "sample" is used in its broadest sense. A sample suspected of
containing MDDT,
nucleic acids encoding MDDT, or fragments thereof may comprise a bodily fluid;
an extract from a
cell, clu-omosome, organelle, or membrane isolated from a cell; a cell;
genomic DNA, RNA, or
cDNA, in solution or bound to a substrate; a tissue; a tissue print; etc.
The terms "specific binding" and "specifically binding" refer to that
interaction between a
protein or peptide and an aganist, an antibody, an antagonist, a small
molecule, or any natural or
synthetic binding composition. The interaction is dependent upon the presence
of a particular
structure of the protein, e.g., the antigenic determinant or epitope,
recognized by the binding
molecule. For example, if an antibody is specific for epitope "A," the
presence of a polypeptide
comprising the epitope A, or the presence of free unlabeled A, in a reaction
containing free labeled A
and the antibody will reduce the amount of labeled A that binds to the
antibody.
The term "substantially purified" refers to nucleic acid or amino acid
sequences that are
removed from their natural environment and are isolated or separated, and are
at least about 60% free,
preferably at least about 75% free, and most preferably at least about 90%
free from other
components with which they are naturally associated.
A "substitution" refers to the replacement of one or more amino acid residues
or nucleotides
by different amino acid residues or nucleotides, respectively.
"Substrate" refers to any suitable rigid or semi-rigid support including
membranes, filters,
chips, slides, wafers, fibers, magnetic or nonmagnetic beads, gels, tubing,
plates, polymers,
microparticles and capillaries. The substrate can have a variety of surface
forms, such as wells,
trenches, pins, channels and pores, to which polynucleotides or polypeptides
are bound.
A "transcript image" or "expression profile" refers to the collective pattern
of gene
expression by a particular cell type or tissue under given conditions at a
given time.
"Transformation" describes a process by which exogenous DNA is introduced into
a recipient
cell. Transformation may occur under natural or artificial conditions
according to various methods
well known in the art, and may rely on any known method for the insertion of
foreign nucleic acid
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sequences into a prokaryotic or eukaryotic host cell. The method for
transformation is selected based
on the type of host cell being transformed and may include, but is not limited
to, bacteriophage or
viral infection, electroporation, heat shock, lipofection, and particle
bombardment. The term
"transformed cells" includes stably transformed cells in which the inserted
DNA is capable of
replication either as an autonomously replicating plasmid or as part of the
host chromosome, as well
as transiently transformed cells which express the inserted DNA or RNA for
limited periods of time.
A "transgenic organism," as used herein, is any organism, including but not
limited to
animals and plants, in which one or more of the cells of the organism contains
heterologous nucleic
acid introduced by way of human intervention, such as by transgenic techniques
well known in the
art. The nucleic acid is introduced into the cell, directly or indirectly by
introduction into a precursor
of the cell, by way of deliberate genetic manipulation, such as by
microinjection or by infection with
a recombinant virus. In another embodiment, the nucleic acid can be introduced
by infection with a
recombinant viral vector, such as a lentiviral vector (Lois, C. et al. (2002)
Science 295:868-872). The
term genetic manipulation does not include classical cross-breeding, or izz
vitro fertilization, but
rather is directed to the introduction of a recombinant DNA molecule. The
transgenic organisms
contemplated in accordance with the present invention include bacteria,
cyanobacteria, fungi, plants
and animals. The isolated DNA of the present invention can be introduced into
the host by methods
known in the art, for example infection, transfection, transformation or
transconjugation. Techniques
for transferring the DNA of the present invention into such organisms are
widely known and
provided in references such as Sambrook and Russell (supra).
A "variant" of a particular nucleic acid sequence is defined as a nucleic acid
sequence having
at least 40% sequence identity to the particular nucleic acid sequence over a
certain length of one of
the nucleic acid sequences using blastn with the "BLAST 2 Sequences" tool
Version 2Ø9 (May-07-
1999) set at default parameters. Such a pair of nucleic acids may show, for
example, at least 50%, at
least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least
91%, at least 92%, at least
93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or
at least 99% or greater
sequence identity over a certain defined length. A variant may be described
as, for example, an
"allelic" (as defined above), "splice," "species," or "polymorphic" variant. A
splice variant may have
significant identity to a reference molecule, but will generally have a
greater or lesser number of
polynucleotides due to alternate splicing of exons during mRNA processing. The
corresponding
polypeptide may possess additional functional domains or lack domains that are
present in the
reference molecule. Species variants are polynucleotides that vary from one
species to another. The
resulting polypeptides will generally have significant amino acid identity
relative to each other. A
polymorphic variant is a variation in the polynucleotide sequence of a
particular gene between
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individuals of a given species. Polymorphic variants also may encompass
"single nucleotide
polymorphisms" (SNPs) in which the polynucleotide sequence varies by one
nucleotide base. The
presence of SNPs may be indicative of, for example, a certain population, a
disease state, or a
propensity for a disease state.
A "variant" of a particular polypeptide sequence is defined as a polypeptide
sequence having
at least 40% sequence identity or sequence similarity to the particular
polypeptide sequence over a
certain length of one of the polypeptide sequences using blastp with the
"BLAST 2 Sequences" tool
Version 2Ø9 (May-07-1999) set at default parameters. Such a pair of
polypeptides may show, for
example, at least 50%, at least 60%, at least 70%, at least 80%, at least 85%,
at least 90%, at least
91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at
least 97%, at least 98%,
or at least 99% or greater sequence identity or sequence similarity over a
certain defined length of one
of the polypeptides.
THE INVENTION
Various embodiments of the invention include new human molecules for disease
detection
and treatment (MDDT), the polynucleotides encoding MDDT, and the use of these
compositions for
the diagnosis, treatment, or prevention of cell proliferative,
autoimmunelinflammatory,
developmental, and neurological disorders.
Table 1 summarizes the nomenclature for the full length polynucleotide and
polypeptide
embodiments of the invention. Each polynucleotide and its corresponding
polypeptide are correlated
to a single Incyte project identification number (Incyte Project ll~). Each
polypeptide sequence is
denoted by both a polypeptide sequence identification number (Polypeptide SEQ
ID NO:) and an
Incyte polypeptide sequence number (Incyte Polypeptide ID) as shown. Each
polynucleotide
sequence is denoted by both a polynucleotide sequence identification number
(Polynucleotide SEQ
ID NO:) and an Incyte polynucleotide consensus sequence number (Incyte
Polynucleotide ~) as
shown. Column 6 shows the Incyte ID numbers of physical, full length clones
corresponding to the
polypeptide and polynucleotide sequences of the invention. The full length
clones encode
polypeptides which have at least 95% sequence identity to the polypeptide
sequences shown in
column 3.
Table 2 shows sequences with homology to the polypeptides of the invention as
identified by
BLAST analysis against the GenBank protein (genpept) database and the PROTEOME
database.
Columns 1 and 2 show the polypeptide sequence identification number
(Polypeptide SEQ >D NO:)
and the corresponding Incyte polypeptide sequence number (Incyte Polypeptide
)D) for polypeptides
of the invention. Column 3 shows the GenBank identification number (GenBank )D
NO:) of the
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nearest GenBank homolog and the PROTEOME database identification numbers
(PROTEOME ID
NO:) of the nearest PROTEOME database homologs. Column 4 shows the probability
scores for the
matches between each polypeptide and its homolog(s). Column 5 shows the
annotation of the
GenBank and PROTEOME database homolog(s) along with relevant citations where
applicable, all of
which are expressly incorporated by reference herein.
Table 3 shows various structural features of the polypeptides of the
invention. Columns 1
and 2 show the polypeptide sequence identification number (SEQ ID NO:) and the
corresponding
Incyte polypeptide sequence number (Incyte Polypeptide ID) for each
polypeptide of the invention.
Column 3 shows the number of amino acid residues in each polypeptide. Column 4
shows potential
phosphorylation sites, and column 5 shows potential glycosylation sites, as
determined by the
MOTIFS program of the GCG sequence analysis software package (Accelrys,
Burlington MA).
Column 6 shows amino acid residues comprising signature sequences, domains,
and motifs. Column
7 shows analytical methods for protein structure/function analysis and in some
cases, searchable
databases to which the analytical methods were applied.
Together, Tables 2 and 3 summarize the properties of polypeptides of the
invention, and these
properties establish that the claimed polypeptides are molecules for disease
detection and treatment.
For example, SEQ 117 N0:26 is 74% identical, from residue Ml to residue P210,
to human
protein similar to WW domain binding protein 2 (GenBank ID g13938601) as
determined by the
Basic Local Alignment Search Tool (BLAST). (See Table 2.) The BLAST
probability score is 2.0e-
73, which indicates the probability of obtaining the observed polypeptide
sequence alignment by
chance. SEQ ID NO:26 is localized to the subcellular region, has binding
protein function, and is a
human WW domain binding protein as determined by BLAST analysis using the
PROTEOME
database. SEQ ID N0:26 also contains a GRAM domain as determined by searching
for statistically
significant matches in the hidden Markov model (HMM)-based PFAM database of
conserved protein
family domains. (See Table 3.) Data from BLAST-PRODOM analysis of the PRODOM
database
provides further corroborative evidence that SEQ ID N0:26 is a WW domain
binding protein.
In another example, SEQ ID N0:40 is 100% identical, from residue M1 to residue
8227 and
from residue 8228 to residue K342, to human NYD-SP6 (GenBank ID g13508446,
residues Ml-
8227 and residues 8267 to K381 respectively) as determined by the Basic Local
Alignment Search
Tool (BLAST). (See Table 2.) The BLAST probability score is 1.1e-195, which
indicates the
probability of obtaining the observed polypeptide sequence alignment by
chance. Data from BLIMPS
analysis provide further corroborative evidence that SEQ ID N0:40 is a PHD-
finger-containing
protein. SEQ ID NO:1-25, SEQ ID N0:27-39, and SEQ ID N0:41-48 were analyzed
and annotated
in a similar manner. The algorithms and parameters for the analysis of SEQ m
NO: l-48 are
CA 02460480 2004-03-15
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described in Table 7.
As shown in Table 4, the full length polynucleotide embodiments were assembled
using
cDNA sequences or coding (exon) sequences derived from genomic DNA, or any
combination of
these two types of sequences. Column 1 lists the polynucleotide sequence
identification number
(Polynucleotide SEQ ID NO:), the corresponding Incyte polynucleotide consensus
sequence number
(Incyte ID) for each polynucleotide of the invention, and the length of each
polynucleotide sequence
in basepairs. Column 2 shows the nucleotide start (5') and stop (3') positions
of the cDNA and/or
genomic sequences used to assemble the full length polynucleotide embodiments,
and of fragments of
the polynucleotides which are useful, for example, in hybridization or
amplification technologies that
identify SEQ ID N0:49-96 or that distinguish between SEQ ID N0:49-96 and
related
polynucleotides.
The polynucleotide fragments described in Column 2 of Table 4 may refer
specifically, for
example, to Incyte cDNAs derived from tissue-specific cDNA libraries or from
pooled cDNA
libraries. Alternatively, the polynucleotide fragments described in column 2
may refer to GenBank
cDNAs or ESTs which contributed to the assembly of the full length
polynucleotides. In addition, the
polynucleotide fragments described in column 2 may identify sequences derived
from the ENSEMBL
(The Sanger Centre, Cambridge, UK) database (i.e., those sequences including
the designation
"ENST"). Alternatively, the polynucleotide fragments described in column 2 may
be derived from
the NCBI RefSeq Nucleotide Sequence Records Database (i. e., those sequences
including the
designation "NM" or "NT") or the NCBI RefSeq Protein Sequence Records (i.e.,
those sequences
including the designation "NP"). Alternatively, the polynucleotide fragments
described in colurrm 2
may refer to assemblages of both cDNA and Genscan-predicted exons brought
together by an "exon
stitching" algorithm. For example, a polynucleotide sequence identified as
FL XXXXXX_N,_Nz_YYYYY_N3 N~ represents a "stitched" sequence in which XXXXXX
is the
identification number of the cluster of sequences to which the algorithm was
applied, and YYYYY is
the number of the prediction generated by the algorithm, and N~,2,3..., if
present, represent specific
exons that may have been manually edited during analysis (See Example V).
Alternatively, the
polynucleotide fragments in column 2 may refer to assemblages of exons brought
together by an
"exon-stretching" algorithm. For example, a polynucleotide sequence identified
as
FLXXXXXX_gA.A.AAA_gBBBBB_1 N is a "stretched" sequence, with XXXXXX being the
Incyte
project identification number, gAAAAA being the GenBank identification number
of the human
genomic sequence to which the "exon-stretching" algorithm was applied, gBBBBB
being the
GenBank identification number or NCBI RefSeq identification number of the
nearest GenBank
protein homolog, and N referring to specific exons (See Example V). In
instances where a RefSeq
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sequence was used as a protein homolog for the "exon-stretching" algorithm, a
RefSeq identifier
(denoted by "NM," "NP," or "NT") may be used in place of the GenBank
identifier (i.e., gBBBBB).
Alternatively, a prefix identifies component sequences that were hand-edited,
predicted from
genomic DNA sequences, or derived from a combination of sequence analysis
methods. The
following Table lists examples of component sequence prefixes and
corresponding sequence analysis
methods associated with the prefixes (see Example IV and Example V).
Prefix Type of analysis and/or examples of programs
GNN, GFG,Exon prediction from genomic sequences using,
for example,
ENST GENSCAN (Stanford University, CA, USA) or
FGENES
(Computer Genomics Group, The Sanger Centre,
Cambridge, UK).
GBI Hand-edited analysis of genomic sequences.
FL Stitched or stretched genomic sequences
(see Example V).
INCY Full length transcript and exon prediction
from mapping of EST
sequences to the genome. Genomic location
and EST composition
data are combined to predict the exons and
resulting transcript.
In some cases, Incyte cDNA coverage redundant with the sequence coverage shown
in Table
4 was obtained to confirm the final consensus polynucleotide sequence, but the
relevant Incyte cDNA
identification numbers are not shown.
Table 5 shows the representative cDNA libraries for those full length
polynucleotides which
were assembled using Incyte cDNA sequences. The representative cDNA library is
the Incyte cDNA
library which is most frequently represented by the Incyte cDNA sequences
which were used to
assemble and confirm the above polynucleotides. The tissues and vectors which
were used to
construct the cDNA libraries shown in Table 5 are described in Table 6.
The invention also encompasses MDDT variants. Various embodiments of MDDT
variants
can have at least about 80%, at least about 90%, or at least about 95% amino
acid sequence identity to
the MDDT amino acid sequence, and can contain at least one functional or
structural characteristic of
MDDT.
Various embodiments also encompass polynucleotides which encode MDDT. In a
particular
embodiment, the invention encompasses a polynucleotide sequence comprising a
sequence selected
from the group consisting of SEQ ID N0:49-96, which encodes MDDT. The
polynucleotide
sequences of SEQ ID N0:49-96, as presented in the Sequence Listing, embrace
the equivalent RNA
sequences, wherein occurrences of the nitrogenous base thymine are replaced
with uracil, and the
sugar backbone is composed of ribose instead of deoxyribose.
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The invention also encompasses variants of a polynucleotide encoding MDDT. In
particular,
such a variant polynucleotide will have at least about 70%, or alternatively
at least about 85%, or
even at least about 95% polynucleotide sequence identity to a polynucleotide
encoding MDDT. A
particular aspect of the invention encompasses a variant of a polynucleotide
comprising a sequence
selected from the group consisting of SEQ ID N0:49-96 which has at least about
70%, or
alternatively at least about 85%, or even at least about 95% polynucleotide
sequence identity to a
nucleic acid sequence selected from the group consisting of SEQ ID N0:49-96.
Any one of the
polynucleotide variants described above can encode a polypeptide which
contains at least one
functional or structural characteristic of MDDT.
In addition, or in the alternative, a polynucleotide variant of the invention
is a splice variant
of a polynucleotide encoding MDDT. A splice variant may have portions which
have significant
sequence identity to a polynucleotide encoding MDDT, but will generally have a
greater or lesser
number of polynucleotides due to additions or deletions of blocks of sequence
arising from alternate
splicing of exons during mRNA processing. A splice variant may have less than
about 70%, or
alternatively less than about 60%, or alternatively less than about 50%
polynucleotide sequence
identity to a polynucleotide encoding MDDT over its entire length; however,
portions of the splice
variant will have at least about 70%, or alternatively at least about 85%, or
alternatively at least about
95%, or alternatively 100% polynucleotide sequence identity to portions of the
polynucleotide
encoding MDDT. For example, a polynucleotide comprising a sequence of SEQ ID
N0:61 is a splice
variant of a polynucleotide comprising a sequence of SEQ ID N0:56. Any one of
the splice variants
described above can encode a polypeptide which contains at least one
functional or structural
characteristic of MDDT.
It will be appreciated by those skilled in the art that as a result of the
degeneracy of the
genetic code, a multitude of polynucleotide sequences encoding MDDT, some
bearing minimal
similarity to the polynucleotide sequences of any known and naturally
occurring gene, may be
produced. Thus, the invention contemplates each and every possible variation
of polynucleotide
sequence that could be made by selecting combinations based on possible codon
choices. These
combinations are made in accordance with the standard triplet genetic code as
applied to the
polynucleotide sequence of naturally occurring MDDT, and all such variations
are to be considered as
being specifically disclosed.
Although polynucleotides which encode MDDT and its variants are generally
capable of
hybridizing to polynucleotides encoding naturally occurring MDDT under
appropriately selected
conditions of stringency, it may be advantageous to produce polynucleotides
encoding MDDT or its
derivatives possessing a substantially different codon usage, e.g., inclusion
of non-naturally occurring
~s
CA 02460480 2004-03-15
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codons. Codons may be selected to increase the rate at which expression of the
peptide occurs in a
particular prokaryotic or eukaryotic host in accordance with the frequency
with which particular
codons are utilized by the host. Other reasons for substantially altering the
nucleotide sequence
encoding MDDT and its derivatives without altering the encoded amino acid
sequences include the
production of RNA transcripts having more desirable properties, such as a
greater half-life, than
transcripts produced from the naturally occurring sequence.
The invention also encompasses production of polynucleotides which encode MDDT
and
MDDT derivatives, or fragments thereof, entirely by synthetic chemistry. After
production, the
synthetic polynucleotide may be inserted into any of the many available
expression vectors and cell
systems using reagents well known in the art. Moreover, synthetic chemistry
may be used to
introduce mutations into a polynucleotide encoding MDDT or any fragment
thereof.
Embodiments of the invention can also include polynucleotides that are capable
of
hybridizing to the claimed polynucleotides, and, in particular, to those
having the sequences shown in
SEQ ID N0:49-96 and fragments thereof, under various conditions of stringency
(Wahl, G.M. and
S.L. Berger (1987) Methods Enzymol. 152:399-407; Kimmel, A.R. (1987) Methods
Enzymol.
152:507-511). Hybridization conditions, including annealing and wash
conditions, are described in
"Definitions."
Methods for DNA sequencing are well known in the art and may be used to
practice any of
the embodiments of the invention. The methods may employ such enzymes as the
Klenow fragment
of DNA polymerise I, SEQUENASE (US Biochemical, Cleveland OH), Taq polymerise
(Applied
Biosystems), thermostable T7 polymerise (Amersham Biosciences, Piscataway NJ),
or combinations
of polymerises and proofreading exonucleases such as those found in the
ELONGASE amplification
system (Invitrogen, Carlsbad CA). Preferably, sequence preparation is
automated with machines such
as the MICROLAB 2200 liquid transfer system (Hamilton, Reno NV), PTC200
thermal cycler (MJ
Research, Watertown MA) and ABI CATALYST 800 thermal cycler (Applied
Biosystems).
Sequencing is then carned out using either the ABI 373 or 377 DNA sequencing
system (Applied
Biosystems), the MEGABACE 1000 DNA sequencing system (Amersham Biosciences),
or other
systems known in the art. The resulting sequences are analyzed using a variety
of algorithms which
are well known in the art (Ausubel et al., supra, ch. 7; Meyers, R.A. (1995)
Molecular Biolo~y and
Biotechnolo~y, Wiley VCH, New York NY, pp. 856-853).
The nucleic acids encoding MDDT may be extended utilizing a partial nucleotide
sequence
and employing various PCR-based methods known in the art to detect upstream
sequences, such as
promoters and regulatory elements. For example, one method which may be
employed,
restriction-site PCR, uses universal and nested primers to amplify unknown
sequence from genomic
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DNA within a cloning vector (Sarkar, G. (1993) PGR Methods Applic. 2:318-322).
Another method,
inverse PCR, uses primers that extend in divergent directions to amplify
unknown sequence from a
circularized template. The template is derived from restriction fragments
comprising a known
genomic locus and surrounding sequences (Triglia, T. et al. (1988) Nucleic
Acids Res. 16:8186). A
third method, capture PCR, involves PCR amplification of DNA fragments
adjacent to known
sequences in human and yeast artificial chromosome DNA (Lagerstrom, M. et al.
(1991) PCR
Methods Applic. 1:l 11-119). In this method, multiple restriction enzyme
digestions and ligations
may be used to insert an engineered double-stranded sequence into a region of
unknown sequence
before performing PCR. Other methods which may be used to retrieve unknown
sequences are
known in the art (Parker, J.D. et al. (1991) Nucleic Acids Res. 19:3055-3060).
Additionally, one may
use PCR, nested primers, and PROMOTERFINDER libraries (Clontech, Palo Alto CA)
to walk
genomic DNA. This procedure avoids the need to screen libraries and is useful
in fording intron/exon
junctions. For all PCR-based methods, primers may be designed using
commercially available
software, such as OLIGO 4.06 primer analysis software (National Biosciences,
Plymouth MN) or
another appropriate program, to be about 22 to 30 nucleotides in length, to
have a GC content of
about 50% or more, and to anneal to the template at temperatures of about
68°C to 72°C.
When screening for full length cDNAs, it is preferable to use libraries that
have been
size-selected to include larger cDNAs. In addition, random-primed libraries,
which often include
sequences containing the 5' regions of genes, are preferable for situations in
which an oligo d(T)
library does not yield a full-length cDNA. Genomic libraries may be useful for
extension of sequence
into 5' non-transcribed regulatory regions.
Capillary electrophoresis systems which are commercially available may be used
to analyze
the size or confirm the nucleotide sequence of sequencing or PCR products. In
particular, capillary
sequencing may employ flowable polymers for electrophoretic separation, four
different nucleotide-
specific, laser-stimulated fluorescent dyes, and a charge coupled device
camera for detection of the
emitted wavelengths. Output/light intensity may be converted to electrical
signal using appropriate
software (e.g., GENOTYPER and SEQUENCE NAVIGATOR, Applied Biosystems), and the
entire
process from loading of samples to computer analysis and electronic data
display may be computer
controlled. Capillary electrophoresis is especially preferable for sequencing
small DNA fragments
which may be present in limited amounts in a particular sample.
In another embodiment of the invention, polynucleotides or fragments thereof
which encode
MDDT may be cloned in recombinant DNA molecules that direct expression of
MDDT, or fragments
or functional equivalents thereof, in appropriate host cells. Due to the
inherent degeneracy of the
genetic code, other polynucleotides which encode substantially the same or a
functionally equivalent
CA 02460480 2004-03-15
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polypeptides may be produced and used to express MDDT.
The polynucleotides of the invention can be engineered using methods generally
known in
the art in order to alter MDDT-encoding sequences for a variety of purposes
including, but not
limited to, modification of the cloning, processing, and/or expression of the
gene product. DNA
shuffling by random fragmentation and PCR reassembly of gene fragments and
synthetic
oligonucleotides may be used to engineer the nucleotide sequences. For
example, oligonucleotide-
mediated site-directed mutagenesis may be used to introduce mutations that
create new restriction
sites, alter glycosylation patterns, change codon preference, produce splice
variants, and so forth.
The nucleotides of the present invention may be subjected to DNA shuffling
techniques such
as MOLECULARBREEDING (Maxygen Inc., Santa Clara CA; described in U.S. Patent
No.
5,837,458; Chang, C.-C. et al. (1999) Nat. Biotechnol. 17:793-797; Christians,
F.C. et al. (1999) Nat.
Biotechnol. 17:259-264; and Crameri, A. et al. (1996) Nat. Biotechnol. 14:315-
319) to alter or
improve the biological properties of MDDT, such as its biological or enzymatic
activity or its ability
to bind to other molecules or compounds. DNA shuffling is a process by which a
library of gene
variants is produced using PCR-mediated recombination of gene fragments. The
library is then
subjected to selection or screening procedures that identify those gene
variants with the desired
properties. These preferred variants may then be pooled and further subjected
to recursive rounds of
DNA shuffling and selection/screening. Thus, genetic diversity is created
through "artificial"
breeding and rapid molecular evolution. For example, fragments of a single
gene containing random
point mutations may be recombined, screened, and then reshuffled until the
desired properties are
optimized. Alternatively, fragments of a given gene may be recombined with
fragments of
homologous genes in the same gene family, either from the same or different
species, thereby
maximizing the genetic diversity of multiple naturally occurring genes in a
directed and controllable
manner.
In another embodiment, polynucleotides encoding MDDT may be synthesized, in
whole or in
part, using one or more chemical methods well known in the art (Caruthers,
M.H. et al. ( 1980)
Nucleic Acids Symp. Ser. 7:215-223; Horn, T. et al. (1980) Nucleic Acids Symp.
Ser. 7:225-232).
Alternatively, MDDT itself or a fragment thereof may be synthesized using
chemical methods known
in the art. For example, peptide synthesis can be performed using various
solution-phase or
solid-phase techniques (Creighton, T. (1984) Proteins, Structures and
Molecular Properties, WH
Freeman, New York NY, pp. 55-60; Roberge, J.Y. et al. (1995) Science 269:202-
204). Automated
synthesis may be achieved using the ABI 431A peptide synthesizer (Applied
Biosystems).
Additionally, the amino acid sequence of MDDT, or any part thereof, may be
altered during direct
synthesis and/or combined with sequences from other proteins, or any part
thereof, to produce a
31
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variant polypeptide or a polypeptide having a sequence of a naturally
occurring polypeptide.
The peptide may be substantially purified by preparative high performance
liquid
chromatography (Chiez, R.M. and F.Z. Regnier (1990) Methods Enzymol. 182:392-
421). The
composition of the synthetic peptides may be confirmed by amino acid analysis
or by sequencing
(Creighton, supra, pp. 28-53).
In order to express a biologically active MDDT, the polynucleotides encoding
MDDT or
derivatives thereof may be inserted into an appropriate expression vector,
i.e., a vector which contains
the necessary elements for transcriptional and translational control of the
inserted coding sequence in
a suitable host. These elements include regulatory sequences, such as
enhancers, constitutive and
inducible promoters, and 5' and 3' untranslated regions in the vector and in
polynucleotides encoding
MDDT. Such elements may vary in their strength and specificity. Specific
initiation signals may
also be used to achieve more efficient translation of polynucleotides encoding
MDDT. Such signals
include the ATG initiation codon and adjacent sequences, e.g. the Kozak
sequence. In cases where a
polynucleotide sequence encoding MDDT and its initiation codon and upstream
regulatory sequences
are inserted into the appropriate expression vector, no additional
transcriptional or translational
control signals may be needed. However, in cases where only coding sequence,
or a fragment
thereof, is inserted, exogenous translational control signals including an in-
frame ATG initiation
codon should be provided by the vector. Exogenous translational elements and
initiation codons may
be of various origins, both natural and synthetic. The efficiency of
expression may be enhanced by
the inclusion of enhancers appropriate for the particular host cell system
used (Scharf, D. et al. (1994)
Results Probl. Cell Differ. 20:125-162).
Methods which are well known to those skilled in the art may be used to
construct expression
vectors containing polynucleotides encoding MDDT and appropriate
transcriptional and translational
control elements. These methods include in vitro recombinant DNA techniques,
synthetic techniques,
and irz vivo genetic recombination (Sambrook and Russell, supra, ch. 1-4, and
8; Ausubel et al.,
supra, ch. l, 3, and 15).
A variety of expression vector/host systems may be utilized to contain and
express
polynucleotides encoding MDDT. These include, but are not limited to,
microorganisms such as
bacteria transformed with recombinant bacteriophage, plasmid, or cosmid DNA
expression vectors;
yeast transformed with yeast expression vectors; insect cell systems infected
with viral expression
vectors (e.g., baculovirus); plant cell systems transformed with viral
expression vectors (e.g.,
cauliflower mosaic virus, CaMV, or tobacco mosaic virus, TMV) or with
bacterial expression vectors
(e.g., Ti or pBR322 plasmids); or animal cell systems (Sambrook and Russell,
supra; Ausubel et al.,
supra; Van Heeke, G. and S.M. Schuster (1989) J. Biol. Chem. 264:5503-5509;
Engelhard, E.K. et al.
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(1994) Proc. Natl. Acad. Sci. USA 91:3224-3227; Sandig, V. et al. (1996) Hum.
Gene Ther. 7:1937-
1945; Takamatsu, N. (1987) EMBO J. 6:307-311; The McGraw Hill Yearbook of
Science and
Technolo~y (1992) McGraw Hill, New York NY, pp. 191-196; Logan, J. and T.
Shenk (1984) Proc.
Natl. Acad. Sci. USA 81:3655-3659; Harrington, J.J. et al. (1997) Nat. Genet.
15:345-355).
Expression vectors derived from retroviruses, adenoviruses, or herpes or
vaccinia viruses, or from
various bacterial plasmids, may be used for delivery of polynucleotides to the
targeted organ, tissue,
or cell population (Di Nicola, M. et al. (1998) Cancer Gen. Ther. 5:350-356;
Yu, M. et al. (1993)
Proc. Natl. Acad. Sci. USA 90:6340-6344; Butler, R.M. et al. (1985) Nature
317:813-815; McGregor,
D.P. et al. (1994) Mol. Immunol. 31:219-226; Verma, LM. and N. Somia (1997)
Nature 389:239-
242). The invention is not limited by the host cell employed.
In bacterial systems, a number of cloning and expression vectors may be
selected depending
upon the use intended for polynucleotides encoding MDDT. For example, routine
cloning,
subcloning, and propagation of polynucleotides encoding MDDT can be achieved
using a
multifunctional E. coli vector such as PBLUESCRIPT (Stratagene, La Jolla CA)
or PSPORTl
plasmid (Invitrogen). Ligation of polynucleotides encoding MDDT into the
vector's multiple cloning
site disrupts the lacZ gene, allowing a colorimetric screening procedure for
identification of
transformed bacteria containing recombinant molecules. In addition, these
vectors may be useful for
in vitro transcription, dideoxy sequencing, single strand rescue with helper
phage, and creation of
nested deletions in the cloned sequence (Van Heeke, G. and S.M. Schuster
(1989) J. Biol. Chem.
264:5503-5509). When large quantities of MDDT are needed, e.g. for the
production of antibodies,
vectors which direct high level expression of MDDT may be used. For example,
vectors containing
the strong, inducible SP6 or T7 bacteriophage promoter may be used.
Yeast expression systems may be used for production of MDDT. A number of
vectors
containing constitutive or inducible promoters, such as alpha factor, alcohol
oxidase, and PGH
promoters, may be used in the yeast Saccharomyces cerevisiae or Piclzia
Pastoris. In addition, such
vectors direct either the secretion or intracellular retention of expressed
proteins and enable
integration of foreign polynucleotide sequences into the host genome for
stable propagation (Ausubel
et al., supra; Bitter, G.A. et al. (1987) Methods Enzymol. 153:516-544;
Scorer, C.A. et al. (1994)
Bio/Technology 12:181-184).
Plant systems may also be used for expression of MDDT. Transcription of
polynucleotides
encoding MDDT may be driven by viral promoters, e.g., the 35S and 19S
promoters of CaMV used
alone or in combination with the omega leader sequence from TMV (Takamatsu, N.
(1987) EMBO J.
6:307-311). Alternatively, plant promoters such as the small subunit of
RUBISCO or heat shock
promoters may be used (Coruzzi, G. et al. (1984) EMBO J. 3:1671-1680; Brogue,
R. et al. (1984)
33
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Science 224:838-843; Winter, J. et al. (1991) Results Probl. Cell Differ.
17:85-105). These
constructs can be introduced into plant cells by direct DNA transformation or
pathogen-mediated
transfection (The McGraw Hill Yearbook of Science and Technolo~y (1992) McGraw
Hill, New
York NY, pp. 191-196).
In mammalian cells, a number of viral-based expression systems may be
utilized. In cases
where an adenovirus is used as an expression vector, polynucleotides encoding
MDDT may be ligated
into an adenovirus transcriptionltranslation complex consisting of the late
promoter and tripartite
leader sequence. Insertion in a non-essential El or E3 region of the viral
genome may be used to
obtain infective virus which expresses MDDT in host cells (Logan, J. and T.
Shenk (1984) Proc. Natl.
Acad. Sci. USA 81:3655-3659). In addition, transcription enhancers, such as
the Rous sarcoma virus
(RSV) enhancer, may be used to increase expression in mammalian host cells.
SV40 or EBV-based
vectors may also be used for high-level protein expression.
Human artificial chromosomes (HACs) may also be employed to deliver larger
fragments of
DNA than can be contained in and expressed from a plasmid. HACs of about 6 kb
to 10 Mb are
constructed and delivered via conventional delivery methods (liposomes,
polycationic amino
polymers, or vesicles) for therapeutic purposes (Harrington, J.J. et al.
(1997) Nat. Genet. 15:345-355).
For long term production of recombinant proteins in mammalian systems, stable
expression
of MDDT in cell lines is preferred. For example, polynucleotides encoding MDDT
can be
transformed into cell lines using expression vectors which may contain viral
origins of replication
and/or endogenous expression elements and a selectable marker gene on the same
or on a separate
vector. Following the introduction of the vector, cells may be allowed to grow
for about 1 to 2 days
in enriched media before being switched to selective media. The purpose of the
selectable marker is
to confer resistance to a selective agent, and its presence allows growth and
recovery of cells which
successfully express the introduced sequences. Resistant clones of stably
transformed cells may be
propagated using tissue culture techniques appropriate to the cell type.
Any number of selection systems may be used to recover transformed cell lines.
These
include, but are not limited to, the herpes simplex virus thymidine leinase
and adenine
phosphoribosyltransferase genes, for use in tk- and apr cells, respectively
(Wigler, M. et al. (1977)
Cell 11:223-232; Lowy, I. et al. (1980) Cell 22:817-823). Also,
antimetabolite, antibiotic, or
herbicide resistance can be used as the basis for selection. For example,
~lhfr confers resistance to
methotrexate; rzeo confers resistance to the aminoglycosides neomycin and G-
418; and als and pat
confer resistance to chlorsulfuron and phosphinotricin acetyltransferase,
respectively (Wigler, M. et
al. (1980) Proc. Natl. Acad. Sci. USA 77:3567-3570; Colbere-Garapin, F. et al.
(1981) J. Mol. Biol.
150:1-14). Additional selectable genes have been described, e.g., trpB and
IzisD, which alter cellular
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CA 02460480 2004-03-15
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requirements for metabolites (Hartman, S.C. and R.C. Mulligan (1988) Proc.
Natl. Acad. Sci. USA
85:8047-8051). Visible markers, e.g., anthocyanins, green fluorescent proteins
(GFP; Clontech), (3-
glucuronidase and its substrate ~3-glucuronide, or luciferase and its
substrate luciferin may be used.
These markers can be used not only to identify transformants, but also to
quantify the amount of
transient or stable protein expression attributable to a specific vector
system (Rhodes, C.A. (1995)
Methods Mol. Biol. 55:121-131).
Although the presence/absence of marker gene expression suggests that the gene
of interest is
also present, the presence and expression of the gene may need to be
confirmed. For example, if the
sequence encoding MDDT is inserted within a marker gene sequence, transformed
cells containing
polynucleotides encoding MDDT can be identified by the absence of marker gene
function.
Alternatively, a marker gene can be placed in tandem with a sequence encoding
MDDT under the
control of a single promoter. Expression of the marker gene in response to
induction or selection
usually indicates expression of the tandem gene as well.
In general, host cells that contain the polynucleotide encoding MDDT and that
express
MDDT may be identified by a variety of procedures known to those of skill in
the art. These
procedures include, but are not limited to, DNA-DNA or DNA-RNA hybridizations,
PCR
amplification, and protein bioassay or immunoassay techniques which include
membrane, solution, or
chip based technologies for the detection and/or quantification of nucleic
acid or protein sequences.
hnmunological methods for detecting and measuring the expression of MDDT using
either
specific polyclonal or monoclonal antibodies are known in the art. Examples of
such techniques
include enzyme-linked immunosorbent assays (ELISAs), radioimmunoassays (RIAs),
and
fluorescence activated cell sorting (FAGS). A two-site, monoclonal-based
immunoassay utilizing
monoclonal antibodies reactive to two non-interfering epitopes on MDDT is
preferred, but a
competitive binding assay may be employed. These and other assays are well
known in the art
(Hampton, R. et al. (1990) Serological Methods, a Laboratory Manual, APS
Press, St. Paul MN, Sect.
IV; Coligan, J.E. et al. (1997) Current Protocols in Immunology, Greene Pub.
Associates and Wiley-
Interscience, New York NY; Pound, J.D. (1998) Immunochemical Protocols, Humana
Press, Totowa
NJ).
A wide variety of labels and conjugation techniques are known by those skilled
in the art and
may be used in various nucleic acid and amino acid assays. Means for producing
labeled
hybridization or PCR probes for detecting sequences related to polynucleotides
encoding MDDT
include oligolabeling, nick translation, end-labeling, or PCR amplification
using a labeled nucleotide.
Alternatively, polynucleotides encoding MDDT, or any fragments thereof, may be
cloned into a
vector for the production of an mRNA probe. Such vectors are known in the art,
are commercially
CA 02460480 2004-03-15
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available, and may be used to synthesize RNA probes in vitro by addition of an
appropriate RNA
polymerise such as T7, T3, or SPG and labeled nucleotides. These procedures
may be conducted
using a variety of commercially available kits, such as those provided by
Amersham Biosciences,
Promega (Madison WI), and US Biochemical. Suitable reporter molecules or
labels which may be
used for ease of detection include radionuclides, enzymes, fluorescent,
chemiluminescent, or
chromogenic agents, as well as substrates, cofactors, inhibitors, magnetic
particles, and the like.
Host cells transformed with polynucleotides encoding MDDT may be cultured
under
conditions suitable for the expression and recovery of the protein from cell
culture. The protein
produced by a transformed cell may be secreted or retained intracellularly
depending on the sequence
and/or the vector used. As will be understood by those of skill in the art,
expression vectors
containing polynucleotides which encode MDDT may be designed to contain signal
sequences which
direct secretion of MDDT through a prokaryotic or eukaryotic cell membrane.
In addition, a host cell strain may be chosen for its ability to modulate
expression of the
inserted polynucleotides or to process the expressed protein in the desired
fashion. Such
modifications of the polypeptide include, but are not limited to, acetylation,
carboxylation,
glycosylation, phosphorylation, lipidation, and acylation. Post-translational
processing which cleaves
a "prepro" or "pro" form of the protein may also be used to specify protein
targeting, folding, and/or
activity. Different host cells which have specific cellular machinery and
characteristic mechanisms
for post-translational activities (e.g., CHO, HeLa, MDCK, HEK293, and WI38)
are available from the
American Type Culture Collection (ATCC, Mantissas VA) and may be chosen to
ensure the correct
modification and processing of the foreign protein.
In another embodiment of the invention, natural, modified, or recombinant
polynucleotides
encoding MDDT may be ligated to a heterologous sequence resulting in
translation of a fusion
protein in any of the aforementioned host systems. For example, a chimeric
MDDT protein
containing a heterologous moiety that can be recognized by a commercially
available antibody may
facilitate the screening of peptide libraries for inhibitors of MDDT activity.
Heterologous protein and
peptide moieties may also facilitate purification of fusion proteins using
commercially available
affinity matrices. Such moieties include, but are not limited to, glutathione
S-transferase (GST),
maltose binding protein (MBP), thioredoxin (Trx), calmodulin binding peptide
(CBP), 6-His, FLAG,
c-nryc, and hemagglutinin (HA). GST, MBP, Trx, CBP, and 6-His enable
purification of their
cognate fusion proteins on immobilized glutathione, maltose, phenylarsine
oxide, calmodulin, and
metal-chelate resins, respectively. FLAG, c-nzyc, and hemagglutinin (HA)
enable immunoaffinity
purification of fusion proteins using commercially available monoclonal and
polyclonal antibodies
that specifically recognize these epitope tags. A fusion protein may also be
engineered to contain a
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proteolytic cleavage site located between the MDDT encoding sequence and the
heterologous protein
sequence, so that MDDT may be cleaved away from the heterologous moiety
following purification.
Methods for fusion protein expression and purification are discussed in
Ausubel et al. (supra, ch. 10
and 16). A variety of commercially available kits may also be used to
facilitate expression and
purification of fusion proteins.
In another embodiment, synthesis of radiolabeled MDDT may be achieved in vitro
using the
TNT rabbit reticulocyte lysate or wheat germ extract system (Promega). These
systems couple
transcription and translation of protein-coding sequences operably associated
with the T7, T3, or SP6
promoters. Translation takes place in the presence of a radiolabeled amino
acid precursor, for
example, 35S-methionine.
MDDT, fragments of MDDT, or variants of MDDT may be used to screen for
compounds
that specifically bind to MDDT. One or more test compounds may be screened for
specific binding
to MDDT. In various embodiments, 1, 2, 3, 4, 5, 10, 20, 50, 100, or 200 test
compounds can be
screened for specific binding to MDDT. Examples of test compounds can include
antibodies,
anticalins, oligonucleotides, proteins (e.g., ligands or receptors), or small
molecules.
In related embodiments, variants of MDDT can be used to screen for binding of
test
compounds, such as antibodies, to MDDT, a variant of MDDT, or a combination of
MDDT andlor
one or more variants MDDT. In an embodiment, a variant of MDDT can be used to
screen for
compounds that bind to a variant of MDDT, but not to MDDT having the exact
sequence of a
sequence of SEQ ID N0:1-48. MDDT variants used to perform such screening can
have a range of
about 50% to about 99% sequence identity to MDDT, with various embodiments
having 60%, 70%,
75%, 80%, 85%, 90%, and 95% sequence identity.
In an embodiment, a compound identified in a screen for specific binding to
MDDT can be
closely related to the natural ligand of MDDT, e.g., a ligand or fragment
thereof, a natural substrate, a
structural or functional mimetic, or a natural binding partner (Coligan, J.E.
et al. (1991) Current
Protocols in Immunolo~Y 1(2):Chapter 5). In another embodiment, the compound
thus identified can
be a natural ligand of a receptor MDDT (Howard, A.D. et al. (2001) Trends
Pharmacol. Sci.22:132-
140; Wise, A. et al. (2002) Drug Discovery Today 7:235-246).
In other embodiments, a compound identified in a screen for specific binding
to MDDT can
be closely related to the natural receptor to which MDDT binds, at least a
fragment of the receptor, or
a fragment of the receptor including all or a portion of the ligand binding
site or binding pocket. For
example, the compound may be a receptor for MDDT which is capable of
propagating a signal, or a
decoy receptor for MDDT which is not capable of propagating a signal
(Ashkenazi, A. and V.M.
Divit (1999) Curr. Opin. Cell Biol. 11:255-260; Mantovani, A. et al. (2001)
Trends Immunol. 22:328-
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336). The compound can be rationally designed using known techniques. Examples
of such
techniques include those used to construct the compound etanercept (ENBREL;
Amgen Inc.,
Thousand Oaks CA), which is efficacious for treating rheumatoid arthritis in
humans. Etanercept is
an engineered p75 tumor necrosis factor (TNF) receptor dimer linked to the Fc
portion of human IgG 1
(Taylor, P.C. et al. (2001) Curr. Opin. Immunol. 13:611-616).
In one embodiment, two or more antibodies having similar or, alternatively,
different
specificities can be screened for specific binding to MDDT, fragments of MDDT,
or variants of
MDDT. The binding specificity of the antibodies thus screened can thereby be
selected to identify
particular fragments or variants of MDDT. In one embodiment, an antibody can
be selected such that
its binding specificity allows for preferential identification of specific
fragments or variants of
MDDT. In another embodiment, an antibody can be selected such that its binding
specificity allows
for preferential diagnosis of a specific disease or condition having
increased, decreased, or otherwise
abnormal production of MDDT.
In an embodiment, anticalins can be screened for specific binding to MDDT,
fragments of
MDDT, or variants of MDDT. Anticalins are ligand-binding proteins that have
been constructed
based on a lipocalin scaffold (Weiss, G.A. and H.B. Lowman (2000) Chem. Biol.
7:8177-8184;
Skerra, A. (2001) J. Biotechnol. 74:257-275). The protein architecture of
lipocalins can include a
beta-barrel having eight antiparallel beta-strands, which supports four loops
at its open end. These
loops form the natural ligand-binding site of the lipocalins, a site which can
be re-engineered ifi vitro
by amino acid substitutions to impart novel binding specificities. The amino
acid substitutions can be
made using methods known in the art or described herein, and can include
conservative substitutions
(e.g., substitutions that do not alter binding specificity) or substitutions
that modestly, moderately, or
significantly alter binding specificity.
In one embodiment, screening for compounds which specifically bind to,
stimulate, or inhibit
MDDT involves producing appropriate cells which express MDDT, either as a
secreted protein or on
the cell membrane. Preferred cells can include cells from mammals, yeast,
Drosophila, or E. coli.
Cells expressing MDDT or cell membrane fractions which contain MDDT are then
contacted with a
test compound and binding, stimulation, or inhibition of activity of either
MDDT or the compound is
analyzed.
An assay may simply test binding of a test compound to the polypeptide,
wherein binding is
detected by a fluorophore, radioisotope, enzyme conjugate, or other detectable
label. For example,
the assay may comprise the steps of combining at least one test compound with
MDDT, either in
solution or affixed to a solid support, and detecting the binding of MDDT to
the compound.
Alternatively, the assay may detect or measure binding of a test compound in
the presence of a
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labeled competitor. Additionally, the assay may be carried out using cell-free
preparations, chemical
libraries, or natural product mixtures, and the test compounds) may be free in
solution or affixed to a
solid support.
An assay can be used to assess the ability of a compound to bind to its
natural ligand andlor
to inhibit the binding of its natural ligand to its natural receptors.
Examples of such assays include
radio-labeling assays such as those described in U.S. Patent No. 5,914,236 and
U.S. Patent No.
6,372,724. In a related embodiment, one or more amino acid substitutions can
be introduced into a
polypeptide compound (such as a receptor) to improve or alter its ability to
bind to its natural ligands
(Matthews, D.J. and J.A. Wells. (1994) Chem. Biol. 1:25-30). In another
related embodiment, one or
more amino acid substitutions can be introduced into a polypeptide compound
(such as a ligand) to
improve or alter its ability to bind to its natural receptors (Cunningham,
B.C. and J.A. Wells (1991)
Proc. Natl. Acad. Sci. USA 88:3407-3411; Lowman, H.B. et al. (1991) J. Biol.
Chem. 266:10982-
10988).
MDDT, fragments of MDDT, or variants of MDDT may be used to screen for
compounds
that modulate the activity of MDDT. Such compounds may include agonists,
antagonists, or partial
or inverse agonists. In one embodiment, an assay is performed under conditions
permissive for
MDDT activity, wherein MDDT is combined with at least one test compound, and
the activity of
MDDT in the presence of a test compound is compared with the activity of MDDT
in the absence of
the test compound. A change in the activity of MDDT in the presence of the
test compound is
indicative of a compound that modulates the activity of MDDT. Alternatively, a
test compound is
combined with an in vitro or cell-free system comprising MDDT under conditions
suitable for MDDT
activity, and the assay is performed. In either of these assays, a test
compound which modulates the
activity of MDDT may do so indirectly and need not come in direct contact with
the test compound.
At least one and up to a plurality of test compounds may be screened.
In another embodiment, polynucleotides encoding MDDT or their mammalian
homologs may
be "knocked out" in an animal model system using homologous recombination in
embryonic stem
(ES) cells. Such techniques are well known in the art and are useful for the
generation of animal
models of human disease (see, e.g., U.S. Patent No. 5,175,383 and U.S. Patent
No. 5,767,337). For
example, mouse ES cells, such as the mouse 129/SvJ cell line, are derived from
the early mouse
embryo and grown in culture. The ES cells are transformed with a vector
containing the gene of
interest disrupted by a marker gene, e.g., the neomycin phosphotransferase
gene (fieo; Capecchi, M.R.
(1989) Science 244:1288-1292). The vector integrates into the corresponding
region of the host
genome by homologous recombination. Alternatively, homologous recombination
takes place using
the Cre-loxP system to knockout a gene of interest in a tissue- or
developmental stage-specific
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manner (Marth, J.D. (1996) Clin. Invest. 97:1999-2002; Wagner, K.U. et al.
(1997) Nucleic Acids
Res. 25:4323-4330). Transformed ES cells are identified and microinjected into
mouse cell
blastocysts such as those from the C57BL/6 mouse strain. The blastocysts are
surgically transferred
to pseudopregnant dams, and the resulting chimeric progeny are genotyped and
bred to produce
S heterozygous or homozygous strains. Transgenic animals thus generated may be
tested with potential
therapeutic or toxic agents.
Polynucle0tides encoding MDDT may also be manipulated in vitro in ES cells
derived from
human blastocysts. Human ES cells have the potential to differentiate into at
least eight separate cell
lineages including endoderm, mesoderm, and ect0dermal cell types. These cell
lineages differentiate
into, for example, neural cells, hematop0ietic lineages, and cardiomyocytes
(Thomson, J.A. et al.
(1998) Science 282:1145-1147).
Polynucleotides encoding MDDT can also be used to create "knockin" humanized
animals
(pigs) or transgenic animals (mice or rats) to model human disease. With
knockin technology, a
region of a polynucleotide encoding MDDT is injected into animal ES cells, and
the injected
sequence integrates into the animal cell genome. Transformed cells are
injected into blastulae, and
the blastulae are implanted as described above. Transgenic progeny or inbred
lines are studied and
treated with potential pharmaceutical agents to obtain information on
treatment of a human disease.
Alternatively, a mammal inbred to overexpress MDDT, e.g., by secreting MDDT in
its milk, may also
serve as a convenient source of that protein (Janne, J. et al. (1998)
Biotechnol. Annu. Rev. 4:55-74).
THERAPEUTICS
Chemical and structural similarity, e.g., in the context of sequences and
motifs, exists
between regions of MDDT and molecules for disease detection and treatment. In
addition, examples
of tissues expressing MDDT can be found in Table 6 and can also be found in
Example XI.
Therefore, MDDT appears to play a role in cell proliferative,
autoimmune/inflammatory,
developmental, and neurological disorders. In the treatment of disorders
associated with increased
MDDT expression or activity, it is desirable to decrease the expression or
activity of MDDT. In the
treatment of disorders associated with decreased MDDT expression or activity,
it is desirable to
increase the expression or activity of MDDT.
Therefore, in one embodiment, MDDT or a fragment or derivative thereof may be
administered to a subject to treat or prevent a disorder associated with
decreased expression or
activity of MDDT. Examples of such disorders include, but are not limited to,
a cell proliferative
disorder such as actinic keratosis, arteriosclerosis, atherosclerosis,
bursitis, cirrhosis, hepatitis, mixed
connective tissue disease (MCTD), myelofibrosis, paroxysmal nocturnal
hemoglobinuria,
polycythemia vera, psoriasis, primary thrombocythemia, and cancers including
adenocarcinoma,
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leukemia, lymphoma, melanoma, myeloma, sarcoma, teratocarcinoma, and, in
particular, cancers of
the adrenal gland, bladder, bone, bone marrow, brain, breast, cervix, gall
bladder, ganglia,
gastrointestinal tract, heart, kidney, liver, lung, muscle, ovary, pancreas,
parathyroid, penis, prostate,
salivary glands, skin, spleen, testis, thymus, thyroid, and uterus; an
autoimmune/inflammatory
disorder such as acquired immunodeficiency syndrome (AIDS), Addison's disease,
adult respiratory
distress syndrome, allergies, ankylosing spondylitis, amyloidosis, anemia,
asthma, atherosclerosis,
autoimmune hemolytic anemia, autoimmune thyroiditis, autoimmune
polyendocrinopathy-
candidiasis-ectodermal dystrophy (APECED), bronchitis, cholecystitis, contact
dermatitis, Crohn's
disease, atopic dermatitis, dermatomyositis, diabetes mellitus, emphysema,
episodic lymphopenia
with lymphocytotoxins, erythroblastosis fetalis, erythema nodosum, atrophic
gastritis,
glomerulonephritis, Goodpasture's syndrome, gout, Graves' disease, Hashimoto's
thyroiditis,
hypereosinophilia, irritable bowel syndrome, multiple sclerosis, myasthenia
gravis, myocardial or
pericardial inflammation, osteoarthritis, osteoporosis, pancreatitis,
polymyositis, psoriasis, Reiter's
syndrome, rheumatoid arthritis, scleroderma, Sjogren's syndrome, systemic
anaphylaxis, systemic
lupus erythematosus, systemic sclerosis, thrombocytopenic purpura, ulcerative
colitis, uveitis, Werner
syndrome, complications of cancer, hemodialysis, and extracorporeal
circulation, viral, bacterial,
fungal, parasitic, protozoal, and helminthic infections, and trauma; a
developmental disorder such as
renal tubular acidosis, anemia, Cushing's syndrome, achondroplastic dwarfism,
Duchenne and Becker
muscular dystrophy, epilepsy, gonadal dysgenesis, WAGR syndrome (Wilms' tumor,
aniridia,
genitourinary abnormalities, and mental retardation), Smith-Magenis syndrome,
myelodysplastic
syndrome, hereditary mucoepithelial dysplasia, hereditary keratodermas,
hereditary neuropathies such
as Charcot-Marie-Tooth disease and neurofibromatosis, hypothyroidism,
hydrocephalus, seizure
disorders such as Syndenham s chorea and cerebral palsy, spina bifida,
anencephaly,
craniorachischisis, congenital glaucoma, cataract, and sensorineural hearing
loss; and a neurological
disorder such as epilepsy, ischemic cerebrovascular disease, stroke, cerebral
neoplasms, Alzheimer's
disease, Pick's disease, Huntington's disease, dementia, Parkinson's disease
and other extrapyramidal
disorders, amyotrophic lateral sclerosis and other motor neuron disorders,
progressive neural
muscular atrophy, retinitis pigmentosa, hereditary ataxias, multiple sclerosis
and other demyelinating
diseases, bacterial and viral meningitis, brain abscess, subdural empyema,
epidural abscess,
suppurative intracranial thrombophlebitis, myelitis and radiculitis, viral
central nervous system
disease, prion diseases including kuru, Creutzfeldt-Jakob disease, and
Gerstmann-
Straussler-Scheinker syndrome, fatal familial insomnia, nutritional and
metabolic diseases of the
nervous system, neurofibromatosis, tuberous sclerosis, cerebelloretinal
hemangioblastomatosis,
encephalotrigeminal syndrome, mental retardation and other developmental
disorders of the central
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nervous system including Down syndrome, cerebral palsy, neuroskeletal
disorders, autonomic
nervous system disorders, cranial nerve disorders, spinal cord diseases,
muscular dystrophy and other
neuromuscular disorders, peripheral nervous system disorders, dermatomyositis
and polymyositis,
inherited, metabolic, endocrine, and toxic myopathies, myasthenia gravis,
periodic paralysis, mental
disorders including mood, anxiety, and schizophrenic disorders, seasonal
affective disorder (SAD),
akathesia, amnesia, catatonia, diabetic neuropathy, tardive dyskinesia,
dystonias, paranoid psychoses,
postherpetic neuralgia, Tourette's disorder, progressive supranuclear palsy,
corticobasal degeneration,
and familial frontotemporal dementia.
In another embodiment, a vector capable of expressing MDDT or a fragment or
derivative
thereof may be administered to a subject to treat or prevent a disorder
associated with decreased
expression or activity of MDDT including, but not limited to, those described
above.
In a further embodiment, a composition comprising a substantially purified
MDDT in
conjunction with a suitable pharmaceutical carrier may be administered to a
subject to treat or prevent
a disorder associated with decreased expression or activity of MDDT including,
but not limited to,
those provided above.
In still another embodiment, an agonist which modulates the activity of MDDT
may be
administered to a subject to treat or prevent a disorder associated with
decreased expression or
activity of MDDT including, but not limited to, those listed above.
In a further embodiment, an antagonist of MDDT may be administered to a
subject to treat or
prevent a disorder associated with increased expression or activity of.MDDT.
Examples of such
disorders include, but are not limited to, those cell proliferative,
autoimmunelinflammatory,
developmental, and neurological disorders described above. In one aspect, an
antibody which
specifically binds MDDT may be used directly as an antagonist or indirectly as
a targeting or delivery
mechanism for bringing a pharmaceutical agent to cells or tissues which
express MDDT.
In an additional embodiment, a vector expressing the complement of the
polynucleotide
encoding MDDT may be administered to a subject to treat or prevent a disorder
associated with
increased expression or activity of MDDT including, but not limited to, those
described above.
In other embodiments, any protein, agonist, antagonist, antibody,
complementary sequence,
or vector embodiments may be administered in combination with other
appropriate therapeutic
agents. Selection of the appropriate agents for use in combination therapy may
be made by one of
ordinary skill in the art, according to conventional pharmaceutical
principles. The combination of
therapeutic agents may act synergistically to effect the treatment or
prevention of the various
disorders described above. Using this approach, one may be able to achieve
therapeutic efficacy with
lower dosages of each agent, thus reducing the potential for adverse side
effects.
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An antagonist of MDDT may be produced using methods which are generally known
in the
art. In particular, purified MDDT may be used to produce antibodies or to
screen libraries of
pharmaceutical agents to identify those which specifically bind MDDT.
Antibodies to MDDT may
also be generated using methods that are well known in the art. Such
antibodies may include, but are
not limited to, polyclonal, monoclonal, chimeric, and single chain antibodies,
Fab fragments, and
fragments produced by a Fab expression library. In an embodiment, neutralizing
antibodies (i.e.,e
those which inhibit dimer formation) can be used therapeutically. Single chain
antibodies (e.g., from
camels or llamas) may be potent enzyme inhibitors and may have application in
the design of peptide
mimetics, and in the development of immuno-adsorbents and biosensors
(Muyldermans, S. (2001) J.
Biotechno1.74:277-302).
For the production of antibodies, various hosts including goats, rabbits,
rats, mice, camels,
dromedaries, llamas, humans, and others may be immunized by injection with
MDDT or with any
fragment or oligopeptide thereof which has immunogenic properties. Depending
on the host species,
various adjuvants may be used to increase immunological response. Such
adjuvants include, but are
not limited to, Freund's, mineral gels such as aluminum hydroxide, and surface
active substances such
as lysolecithin, pluronic polyols, polyanions, peptides, oil emulsions, KLH,
and dinitrophenol.
Among adjuvants used in humans, BCG (bacilli Calmette-Guerin) and
Corynebacteriusn ~arvunz are
especially preferable.
It is preferred that the oligopeptides, peptides, or fragments used to induce
antibodies to
MDDT have an amino acid sequence consisting of at least about 5 amino acids,
and generally will
consist of at least about 10 amino acids. It is also preferable that these
oligopeptides, peptides, or
fragments are substantially identical to a portion of the amino acid sequence
of the natural protein.
Short stretches of MDDT amino acids may be fused with those of another
protein, such as KLH, and
antibodies to the chimeric molecule may be produced.
Monoclonal antibodies to MDDT may be prepared using any technique which
provides for
the production of antibody molecules by continuous cell lines in culture.
These include, but are not
limited to, the hybridoma technique, the human B-cell hybridoma technique, and
the EBV-hybridoma
technique (Kohler, G. et al. (1975) Nature 256:495-497; Kozbor, D. et al.
(1985) J. Immunol.
Methods 81:31-42; Cote, R.J. et al. (1983) Proc. Natl. Acad. Sci. USA 80:2026-
2030; Cole, S.P. et al.
(1984) Mol. Cell Biol. 62:109-120).
In addition, techniques developed for the production of "chimeric antibodies,"
such as the
splicing of mouse antibody genes to human antibody genes to obtain a molecule
with appropriate
antigen specificity and biological activity, can be used (Morrison, S.L. et
al. (1984) Proc. Natl. Acad.
Sci. USA 81:6851-6855; Neuberger, M.S. et al. (1984) Nature 312:604-608;
Takeda, S. et al. (1985)
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Nature 314:452-454). Alternatively, techniques described for the production of
single chain
antibodies may be adapted, using methods known in the art, to produce MDDT-
specific single chain
antibodies. Antibodies with related specificity, but of distinct idiotypic
composition, may be
generated by chain shuffling from random combinatorial immunoglobulin
libraries (Burton, D.R.
(1991) Proc. Natl. Acad. Sci. USA 88:10134-10137).
Antibodies may also be produced by inducing in vivo production in the
lymphocyte
population or by screening immunoglobulin libraries or panels of highly
specific binding reagents as
disclosed in the literature (Orlandi, R. et al. (1989) Proc. Natl. Acad. Sci.
USA 86:3833-3837; Winter,
G. et al. (1991) Nature 349:293-299).
Antibody fragments which contain specific binding sites for MDDT may also be
generated.
For example, such fragments include, but are not limited to, F(ab~2 fragments
produced by pepsin
digestion of the antibody molecule and Fab fragments generated by reducing the
disulfide bridges of
the F(ab~2 fragments. Alternatively, Fab expression libraries may be
constructed to allow rapid and
easy identification of monoclonal Fab fragments with the desired specificity
(Huse, W.D. et al. (1989)
Science 246:1275-1281).
Various immunoassays may be used for screening to identify antibodies having
the desired
specificity. Numerous protocols for competitive binding or immunoradiometric
assays using either
polyclonal or monoclonal antibodies with established specificities are well
laiown in the art. Such
immunoassays typically involve the measurement of complex formation between
MDDT and its
specific antibody. A two-site, monoclonal-based immunoassay utilizing
monoclonal antibodies
i
reactive to two non-interfering MDDT epitopes is generally used, but a
competitive binding assay
may also be employed (Pound, supra).
Various methods such as Scatchard analysis in conjunction with
radioimmunoassay
techniques may be used to assess the affinity of antibodies for MDDT. Affinity
is expressed as an
association constant, Ka, which is defined as the molar concentration of MDDT-
antibody complex
divided by the molar concentrations of free antigen and free antibody under
equilibrium conditions.
The Ka determined for a preparation of polyclonal antibodies, which are
heterogeneous in their
affinities for multiple MDDT epitopes, represents the average affinity, or
avidity, of the antibodies
for MDDT. The Ka determined for a preparation of monoclonal antibodies, which
are monospecific
for a particular MDDT epitope, represents a true measure of affinity. High-
affinity antibody
preparations with Ka ranging from about 109 to 10'2 Llmole are preferred for
use in immunoassays in
which the MDDT-antibody complex must withstand rigorous manipulations. Low-
affinity antibody
preparations with Ka ranging from about 106 to 10' L/mole are preferred for
use in
immunopurification and similar procedures which ultimately require
dissociation of MDDT,
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preferably in active form, from the antibody (Catty, D. (1988) Antibodies,
Volume I: A Practical
Auproach, IRL Press, Washington DC; Liddell, J.E. and A. Cryer (1991) A
Practical Guide to
Monoclonal Antibodies, John Wiley & Sons, New York NY).
The titer and avidity of polyclonal antibody preparations may be further
evaluated to
determine the quality and suitability of such preparations for certain
downstream applications. For
example, a polyclonal antibody preparation containing at least 1-2 mg specific
antibody/ml,
preferably 5-10 mg specific antibody/ml, is generally employed in procedures
requiring precipitation
of MDDT-antibody complexes. Procedures for evaluating antibody specificity,
titer, and avidity, and
guidelines for antibody quality and usage in various applications, are
generally available (Catty,
stapra; Coligan et al., supra).
In another embodiment of the invention, polynucleotides encoding MDDT, or any
fragment
or complement thereof, may be used for therapeutic purposes. In one aspect,
modifications of gene
expression can be achieved by designing complementary sequences or antisense
molecules (DNA,
RNA, PNA, or modified oligonucleotides) to the coding or regulatory regions of
the gene encoding
MDDT. Such technology is well known in the art, and antisense oligonucleotides
or larger fragments
can be designed from various locations along the coding or control regions of
sequences encoding
MDDT (Agrawal, S., ed. (1996) Antisense Therapeutics, Humana Press, Totawa
NJ).
In therapeutic use, any gene delivery system suitable for introduction of the
antisense
sequences into appropriate target cells can be used. Antisense sequences can
be delivered
intracellularly in the form of an expression plasmid which, upon
transcription, produces a sequence
complementary to at least a portion of the cellular sequence encoding the
target protein (Slater, J.E. et
al. (1998) J. Allergy Clin. Immunol. 102:469-475; Scanlon, K.J. et al. (1995)
9:1288-1296).
Antisense sequences can also be introduced intracellularly through the use of
viral vectors, such as
retrovirus and adeno-associated virus vectors (Miller, A.D. (1990) Blood
76:271; Ausubel et al.,
supra; Uckert, W. and W. Walther (1994) Pharmacol. Ther. 63:323-347). Other
gene delivery
mechanisms include liposome-derived systems, artificial viral envelopes, and
other systems known in
the art (Rossi, J.J. (1995) Br. Med. Bull. 51:217-225; Boado, R.J. et al.
(1998) J. Pharm. Sci.
87:1308-1315; Morris, M.C. et al. (1997) Nucleic Acids Res. 25:2730-2736).
In another embodiment of the invention, polynucleotides encoding MDDT may be
used for
somatic or germline gene therapy. Gene therapy may be performed to (i) correct
a genetic deficiency
(e.g., in the cases of severe combined immunodeficiency (SCID)-Xl disease
characterized by X-
linked inheritance (Cavazzana-Calvo, M. et al. (2000) Science 288:669-672),
severe combined
immunodeficiency syndrome associated with an inherited adenosine deaminase
(ADA) deficiency
(Blaese, R.M. et al. (1995) Science 270:475-480; Bordignon, C. et al. (1995)
Science 270:470-475),
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cystic fibrosis (Zabner, J. et al. (1993) Cel175:207-216; Crystal, R.G. et al.
(1995) Hum. Gene
Therapy 6:643-666; Crystal, R.G. et al. (1995) Hum. Gene Therapy 6:667-703),
thalassamias, familial
hypercholesterolemia, and hemophilia resulting from Factor VIII or Factor IX
deficiencies (Crystal,
R.G. (1995) Science 270:404-410; Verma, LM. and N. Somia (1997) Nature 389:239-
242)), (ii)
express a conditionally lethal gene product (e.g., in the case of cancers
which result from unregulated
cell proliferation), or (iii) express a protein which affords protection
against intracellular parasites
(e.g., against human retroviruses, such as human immunodeficiency virus (HIV)
(Baltimore, D.
(1988) Nature 335:395-396; Poeschla, E. et al. (1996) Proc. Natl. Acad. Sci.
USA 93:11395-11399),
hepatitis B or C virus (HBV, HCV); fungal parasites, such as Cazzdida
albicazzs and Paracoccidioides
brasili.ezzsis; and protozoan parasites such as Plaszzzodiunz falciparum and
Trypazzosozzza cruzi). In the
case where a genetic deficiency in MDDT expression or regulation causes
disease, the expression of
MDDT from an appropriate population of transduced cells may alleviate the
clinical manifestations
caused by the genetic deficiency.
In a further embodiment of the invention, diseases or disorders caused by
deficiencies in
MDDT are treated by constructing mammalian expression vectors encoding MDDT
and introducing
these vectors by mechanical means into MDDT-deficient cells. Mechanical
transfer technologies for
use with cells izz vivo or ex vitro include (i) direct DNA microinjection into
individual cells, (ii)
ballistic gold particle delivery, (iii) liposome-mediated transfection, (iv)
receptor-mediated gene
transfer, and (v) the use of DNA transposons (Morgan, R.A. and W.F. Anderson
(1993) Annu. Rev.
Biochem. 62:191-217; Ivics, Z. (1997) Cell 91:501-510; Boulay, J.-L. and H.
Recipon (1998) Curr.
Opin. Biotechnol. 9:445-450).
Expression vectors that may be effective for the expression of MDDT include,
but are not
limited to, the PCDNA 3.1, EPITAG, PRCCMV2, PREP, PVAX, PCR2-TOPOTA vectors
(Invitrogen, Carlsbad CA), PCMV-SCRIPT, PCMV-TAG, PEGSH/PERV (Stratagene, La
Jolla CA),
and PTET-OFF, PTET-ON, PTRE2, PTRE2-LUC, PTK-HYG (Clontech, Palo Alto CA).
MDDT
may be expressed using (i) a constitutively active promoter, (e.g., from
cytomegalovirus (CMV),
Rous sarcoma virus (RSV), SV40 virus, thymidine kinase (TK), or (3-actin
genes), (ii) an inducible
promoter (e.g., the tetracycline-regulated promoter (Gossen, M. and H. Bujard
(1992) Proc. Natl.
Acad. Sci. USA 89:5547-5551; Gossen, M. et al. (1995) Science 268:1766-1769;
Rossi, F.M.V. and
H.M. Blau (1998) Curr. Opin. Biotechnol. 9:451-456), commercially available in
the T-REX plasmid
(Invitrogen)); the ecdysone-inducible promoter (available in the plasmids
PVGRXR and PIND;
Invitrogen); the FK506/rapamycin inducible promoter; or the RU486/mifepristone
inducible promoter
(Rossi, F.M.V. and H.M. Blau, supra)), or (iii) a tissue-specific promoter or
the native promoter of
the endogenous gene encoding MDDT from a normal individual.
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Commercially available liposome transformation kits (e.g., the PERFECT LIPID
TRANSFECTION KIT, available from Invitrogen) allow one with ordinary skill in
the art to deliver
polynucleotides to target cells in culture and require minimal effort to
optimize experimental
parameters. In the alternative, transformation is performed using the calcium
phosphate method
(Graham, F.L. and A.J. Eb (1973) Virology 52:456-467), or by electroporation
(Neumann, E. et al.
(1982) EMBO J. 1:841-845). The introduction of DNA to primary cells requires
modification of
these standardized mammalian transfection protocols.
In another embodiment of the invention, diseases or disorders caused by
genetic defects with
respect to MDDT expression are treated by constructing a retrovirus vector
consisting of (i) the
polynucleotide encoding MDDT under the control of an independent promoter or
the retrovirus long
terminal repeat (LTR) promoter, (ii) appropriate RNA packaging signals, and
(iii) a Rev-responsive
element (RRE) along with additional retrovirus cis-acting RNA sequences and
coding sequences
required for efficient vector propagation. Retrovirus vectors (e.g., PFB and
PFSNEO) are
commercially available (Stratagene) and are based on published data (Riviere,
I. et al. (1995) Proc.
Natl. Acad. Sci. USA 92:6733-6737), incorporated by reference herein. The
vector is propagated in
an appropriate vector producing cell line (VPCL) that expresses an envelope
gene with a tropism for
receptors on the target cells or a promiscuous envelope protein such as VSVg
(Armentano, D. et al.
(1987) J. Virol. 61:1647-1650; Bender, M.A. et al. (1987) J. Virol. 61:1639-
1646; Adam, M.A. and
A.D. Miller (1988) J. Virol. 62:3802-3806; Dull, T. et al. (1998) J. Virol.
72:8463-8471; Zufferey, R.
et al. ( 1998) J. Virol. 72:9873-9880). U.S. Patent No. 5,910,434 to Rigg
("Method for obtaining
retrovirus packaging cell lines producing high transducing efficiency
retroviral supernatant")
discloses a method for obtaining retrovirus packaging cell lines and is hereby
incorporated by
reference. Propagation of retrovirus vectors, transduction of a population of
cells (e.g., CD4+ T-
cells), and the return of transduced cells to a patient are procedures well
known to persons skilled in
the art of gene therapy and have been well documented (Ranga, U. et al. (1997)
J. Virol. 71:7020-
7029; Bauer, G. et al. (1997) Blood 89:2259-2267; Bonyhadi, M.L. (1997) J.
Virol. 71:4707-4716;
Ranga, U. et al. (1998) Proc. Natl. Acad. Sci. USA 95:1201-1206; Su, L. (1997)
Blood 89:2283-
2290).
In an embodiment, an adenovirus-based gene therapy delivery system is used to
deliver
polynucleotides encoding MDDT to cells which have one or more genetic
abnormalities with respect
to the expression of MDDT. The construction and packaging of adenovirus-based
vectors are well
known to those with ordinary skill in the art. Replication defective
adenovirus vectors have proven to
be versatile for importing genes encoding immunoregulatory proteins into
intact islets in the pancreas
(Csete, M.E. et al. (1995) Transplantation 27:263-268). Potentially useful
adenoviral vectors are
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described in U.S. Patent No. 5,707,618 to Armentano ("Adenovirus vectors for
gene therapy"),
hereby incorporated by reference. For adenoviral vectors, see also Antinozzi,
P.A. et al. ( 1999; Annu.
Rev. Nutr. 19:511-544) and Verma, LM. and N. Somia (1997; Nature 18:389:239-
242).
In another embodiment, a herpes-based, gene therapy delivery system is used to
deliver
polynucleotides encoding MDDT to target cells which have one or more genetic
abnormalities with
respect to the expression of MDDT. The use of herpes simplex virus (HSV)-based
vectors may be
especially valuable for introducing MDDT to cells of the central nervous
system, for which HSV has
a tropism. The construction and packaging of herpes-based vectors are well
known to those with
ordinary skill in the art. A replication-competent herpes simplex virus (HSV)
type 1-based vector has
been used to deliver a reporter gene to the eyes of primates (Liu, X. et al.
(1999) Exp. Eye Res.
169:385-395). The construction of a HSV-1 virus vector has also been disclosed
in detail in U.S.
Patent No. 5,804,413 to DeLuca ("Herpes simplex virus strains for gene
transfer"), which is hereby
incorporated by reference. U.S. Patent No. 5,804,413 teaches the use of
recombinant HSV d92 which
consists of a genome containing at least one exogenous gene to be transferred
to a cell under the
control of the appropriate promoter for purposes including human gene therapy.
Also taught by this
patent are the construction and use of recombinant HSV strains deleted for
ICP4, ICP27 and ICP22.
For HSV vectors, see also Goins, W.F. et al. (1999; J. Virol. 73:519-532) and
Xu, H. et al. (1994;
Dev. Biol. 163:152-161). The manipulation of cloned herpesvirus sequences, the
generation of
recombinant virus following the transfection of multiple plasmids containing
different segments of
the large herpesvirus genomes, the growth and propagation of herpesvirus, and
the infection of cells
with herpesvirus are techniques well known to those of ordinary skill in the
art.
In another embodiment, an alphavirus (positive, single-stranded RNA virus)
vector is used to
deliver polynucleotides encoding MDDT to target cells. The biology of the
prototypic alphavirus,
Semliki Forest Virus (SFV), has been studied extensively and gene transfer
vectors have been based
on the SFV genome (Garoff, H. and K.-J. Li (1998) Curr. Opin. Biotechnol.
9:464-469). During
alphavirus RNA replication, a subgenomic RNA is generated that normally
encodes the viral capsid
proteins. This subgenomic RNA replicates to higher levels than the full length
genomic RNA,
resulting in the overproduction of capsid proteins relative to the viral
proteins with enzymatic activity
(e.g., protease and polymerase). Similarly, inserting the coding sequence for
MDDT into the
alphavirus genome in place of the capsid-coding region results in the
production of a large number of
MDDT-coding RNAs and the synthesis of high levels of MDDT in vector transduced
cells. While
alphavirus infection is typically associated with cell lysis within a few
days, the ability to establish a
persistent infection in hamster normal kidney cells (BHK-21) with a variant of
Sindbis virus (SIN)
indicates that the lytic replication of alphaviruses can be altered to suit
the needs of the gene therapy
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application (Dryga, S.A. et al. (1997) Virology 228:74-83). The wide host
range of alphaviruses will
allow the introduction of MDDT into a variety of cell types. The specific
transduction of a subset of
cells in a population may require the sorting of cells prior to transduction.
The methods of
manipulating infectious cDNA clones of alphaviruses, performing alphavirus
cDNA and RNA
transfections, and performing alphavirus infections, are well known to those
with ordinary skill in the
art.
Oligonucleotides derived from the transcription initiation site, e.g., between
about positions
-10 and +10 from the start site, may also be employed to inhibit gene
expression. Similarly,
inhibition can be achieved using triple helix base-pairing methodology. Triple
helix pairing is useful
because it causes inhibition of the ability of the double helix to open
sufficiently for the binding of
polymerases, transcription factors, or regulatory molecules. Recent
therapeutic advances using
triplex DNA have been described in the literature (Gee, J.E. et al. (1994) in
Huber, B.E. and B.I. Carr,
Molecular and Immunolo~ic Approaches, Futura Publishing, Mt. Kisco NY, pp. 163-
177). A
complementary sequence or antisense molecule may also be designed to block
translation of mRNA
by preventing the transcript from binding to ribosomes.
Ribozymes, enzymatic RNA molecules, may also be used to catalyze the specific
cleavage of
RNA. The mechanism of ribozyme action involves sequence-specific hybridization
of the ribozyme
molecule to complementary target RNA, followed by endonucleolytic cleavage.
For example,
engineered hammerhead motif ribozyme molecules may specifically and
efficiently catalyze
endonucleolytic cleavage of RNA molecules encoding MDDT.
Specific ribozyme cleavage sites within any potential RNA target are initially
identified by
scanning the target molecule for ribozyme cleavage sites, including the
following sequences: GUA,
GUU, and GUC. Once identified, short RNA sequences of between 15 and 20
ribonucleotides,
corresponding to the region of the target gene containing the cleavage site,
may be evaluated for
secondary structural features which may render the oligonucleotide inoperable.
The suitability of
candidate targets may also be evaluated by testing accessibility to
hybridization with complementary
oligonucleotides using ribonuclease protection assays.
Complementary ribonucleic acid molecules and ribozymes may be prepared by any
method
known in the art for the synthesis of nucleic acid molecules. These include
techniques for chemically
synthesizing oligonucleotides such as solid phase phosphoramidite chemical
synthesis. Alternatively,
RNA molecules may be generated by izz vitro and izz vivo transcription of DNA
molecules encoding
MDDT. Such DNA sequences may be incorporated into a wide variety of vectors
with suitable RNA
polymerase promoters such as T7 or SP6. Alternatively, these cDNA constructs
that synthesize
complementary RNA, constitutively or inducibly, can be introduced into cell
lines, cells, or tissues.
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RNA molecules may be modified to increase intracellular stability and half-
life. Possible
modifications include, but are not limited to, the addition of flanking
sequences at the 5' and/or 3'
ends of the molecule, or the use of phosphorothioate or 2' O-methyl rather
than phosphodiesterase
linkages within the backbone of the molecule. This concept is inherent in the
production of PNAs
and can be extended in all of these molecules by the inclusion of
nontraditional bases such as inosine,
queosine, and wybutosine, as well as, acetyl-, methyl-, thio-, and similarly
modified forms of adenine,
cytidine, guanine, thymine, and uridine which are not as easily recognized by
endogenous
endonucleases.
In other embodiments of the invention, the expression of one or more selected
polynucleotides of the present invention can be altered, inhibited, decreased,
or silenced using RNA
interference (RNAi) or post-transcriptional gene silencing (PTGS) methods
known in the art. RNAi
is a post-transcriptional mode of gene silencing in which double-stranded RNA
(dsRNA) introduced
into a targeted cell specifically suppresses the expression of the homologous
gene (i.e., the gene
bearing the sequence complementary to the dsRNA). This effectively knocks out
or substantially
reduces the expression of the targeted gene. PTGS can also be accomplished by
use of DNA or DNA
fragments as well. RNAi methods are described by Fire, A. et al. (1998; Nature
391:806-811) and
Gura, T. (2000; Nature 404:804-808). PTGS can also be initiated by
introduction of a
complementary segment of DNA into the selected tissue using gene delivery
and/or viral vector
delivery methods described herein or known in the art.
RNAi can be induced in mammalian cells by the use of small interfering RNA
also known as
siRNA. SiRNA are shorter segments of dsRNA (typically about 21 to 23
nucleotides in length) that
result in vivo from cleavage of introduced dsRNA by the action of an
endogenous ribonuclease.
SiRNA appear to be the mediators of the RNAi effect in mammals. The most
effective siRNAs
appear to be 21 nucleotide dsRNAs with 2 nucleotide 3' overhangs. The use of
siRNA for inducing
RNAi in mammalian cells is described by Elbashir, S.M. et al. (2001; Nature
411:494-498).
SiRNA can either be generated indirectly by introduction of dsRNA into the
targeted cell, or
directly by mammalian transfection methods and agents described herein or
known in the art (such as
liposome-mediated transfection, viral vector methods, or other polynucleotide
delivery/introductory
methods). Suitable SiRNAs can be selected by examining a transcript of the
target polynucleotide
(e.g., mRNA) for nucleotide sequences downstream from the AUG start codon and
recording the
occurrence of each nucleotide and the 3' adjacent 19 to 23 nucleotides as
potential siRNA target sites,
with sequences having a 21 nucleotide length being preferred. Regions to be
avoided for target
siRNA sites include the 5' and 3' untranslated regions (UTRs) and regions near
the start codon (within
75 bases), as these may be richer in regulatory protein binding sites. UTR-
binding proteins and/or
CA 02460480 2004-03-15
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translation initiation complexes may interfere with binding of the siRNP
endonuclease complex. The
selected target sites for siRNA can then be compared to the appropriate genome
database (e.g.,
human, etc.) using BLAST or other sequence comparison algorithms known in the
art. Target
sequences with significant homology to other coding sequences can be
eliminated from consideration.
The selected SiRNAs can be produced by chemical synthesis methods known in the
art or by ira vitro
transcription using commercially available methods and kits such as the
SILENCER siRNA
construction kit (Ambion, Austin TX).
In alternative embodiments, long-term gene silencing and/or RNAi effects can
be induced in
selected tissue using expression vectors that continuously express siRNA. This
can be accomplished
using expression vectors that are engineered to express hairpin RNAs (shRNAs)
using methods
known in the art (see, e.g., Brummelkamp, T.R. et al. (2002) Science 296:550-
553; and Paddison, P.J.
et al. (2002) Genes Dev. 16:948-958). In these and related embodiments, shRNAs
can be delivered to
target cells using expression vectors known in the art. An example of a
suitable expression vector for
delivery of siRNA is the PSILENCER1.0-U6 (circular) plasmid (Ambion). Once
delivered to the
target tissue, shRNAs are processed ifa vivo into siRNA-like molecules capable
of carrying out gene-
specific silencing.
In various embodiments, the expression levels of genes targeted by RNAi or
PTGS methods
can be determined by assays for nnRNA and/or protein analysis. Expression
levels of the mRNA of a
targeted gene, can be determined by northern analysis methods using, for
example, the
NORTHERNMAX-GLY kit (Ambion); by microarray methods; by PCR methods; by real
time PCR
methods; and by other RNA/polynucleotide assays known in the art or described
herein. Expression
levels of the protein encoded by the targeted gene can be determined by
Western analysis using
standard techniques known in the art.
An additional embodiment of the invention encompasses a method for screening
for a
compound which is effective in altering expression of a polynucleotide
encoding MDDT.
Compounds which may be effective in altering expression of a specific
polynucleotide may include,
but are not limited to, oligonucleotides, antisense oligonucleotides, triple
helix-forming
oligonucleotides, transcription factors and other polypeptide transcriptional
regulators, and non-
macromolecular chemical entities which are capable of interacting with
specific polynucleotide
sequences. Effective compounds may alter polynucleotide expression by acting
as either inhibitors or
promoters of polynucleotide expression. Thus, in the treatment of disorders
associated with increased
MDDT expression or activity, a compound which specifically inhibits expression
of the
polynucleotide encoding MDDT may be therapeutically useful, and in the
treatment of disorders
associated with decreased MDDT expression or activity, a compound which
specifically promotes
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expression of the polynucleotide encoding MDDT may be therapeutically useful.
In various embodiments, one or more test compounds may be screened for
effectiveness in
altering expression of a specific polynucleotide. A test compound may be
obtained by any method
commonly known in the art, including chenucal modification of a compound known
to be effective in
altering polynucleotide expression; selection from an existing, commercially-
available or proprietary
library of naturally-occurring or non-natural chemical compounds; rational
design of a compound
based on chemical and/or structural properties of the target polynucleotide;
and selection from a
library of chemical compounds created combinatorially or randomly. A sample
comprising a
polynucleotide encoding MDDT is exposed to at least one test compound thus
obtained. The sample
may comprise, for example, an intact or permeabilized cell, or an in vitro
cell-free or reconstituted
biochemical system. Alterations in the expression of a polynucleotide encoding
MDDT are assayed
by any method commonly known in the art. Typically, the expression of a
specific nucleotide is
detected by hybridization with a probe having a nucleotide sequence
complementary to the sequence
of the polynucleotide encoding MDDT. The amount of hybridization may be
quantified, thus
forming the basis for a comparison of the expression of the polynucleotide
both with and without
exposure to one or more test compounds. Detection of a change in the
expression of a polynucleotide
exposed to a test compound indicates that the test compound is effective in
altering the expression of
the polynucleotide. A screen for a compound effective in altering expression
of a specific
polynucleotide can be carried out, for example, using a Schizosacclaaronayces
pomve gene expression
system (Atkins, D. et al. (1999) U.S. Patent No. 5,932,435; Arndt, G.M. et al.
(2000) Nucleic Acids
Res. 28:E15) or a human cell line such as HeLa cell (Clarke, M.L. et al.
(2000) Biochem. Biophys.
Res. Commun. 268:8-13). A particular embodiment of the present invention
involves screening a
combinatorial library of oligonucleotides (such as deoxyribonucleotides,
ribonucleotides, peptide
nucleic acids, and modified oligonucleotides) for antisense activity against a
specific polynucleotide
sequence (Bruice, T.W. et al. (1997) U.S. Patent No. 5,686,242; Bruice, T.W.
et al. (2000) U.S.
Patent No. 6,022,691).
Many methods for introducing vectors into cells or tissues are available and
equally suitable
for use in vivo, irz vitro, and ex vivo. For ex vivo therapy, vectors may be
introduced into stem cells
taken from the patient and clonally propagated for autologous transplant back
into that same patient.
Delivery by transfection, by liposome injections, or by polycatioriic amino
polymers may be achieved
using methods which are well known in the art (Goldman, C.K. et al. (1997)
Nat. Biotechnol. 15:462-
466).
Any of the therapeutic methods described above may be applied to any subject
in need of
such therapy, including, fox example, mammals such as humans, dogs, cats,
cows, horses, rabbits, and
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monkeys.
An additional embodiment of the invention relates to the administration of a
composition
which generally comprises an active ingredient formulated with a
pharmaceutically acceptable
excipient. Excipients may include, for example, sugars, starches, celluloses,
gums, and proteins.
Various formulations are commonly known and are thoroughly discussed in the
latest edition of
Remin~ton's Pharmaceutical Sciences (Maack Publishing, Easton PA). Such
compositions may
consist of MDDT, antibodies to MDDT, and mimetics, agonists, antagonists, or
inhibitors of MDDT.
In various embodiments, the compositions described herein, such as
pharmaceutical
compositions, may be administered by any number of routes including, but not
limited to, oral,
intravenous, intramuscular, intra-arterial, intramedullary, intrathecal,
intraventricular, pulmonary,
transdermal, subcutaneous, intraperitoneal, intranasal, enteral, topical,
sublingual, or rectal means.
Compositions for pulmonary administration may be prepared in liquid or dry
powder form.
These compositions are generally aerosolized immediately prior to inhalation
by the patient. In the
case of small molecules (e.g. traditional low molecular weight organic drugs),
aerosol delivery of
fast-acting formulations is well-known in the art. In the case of
macromolecules (e.g. larger peptides
and proteins), recent developments in the field of pulmonary delivery via the
alveolar region of the
lung have enabled the practical delivery of drugs such as insulin to blood
circulation (see, e.g., Patton,
J.S. et al., U.S. Patent No. 5,997,848). Pulmonary delivery allows
administration without needle
injection, and obviates the need for potentially toxic penetration enhancers.
Compositions suitable for use in the invention include compositions wherein
the active
ingredients are contained in an effective amount to achieve the intended
purpose. The determination
of an effective dose is well within the capability of those skilled in the
art.
Specialized forms of compositions may be prepared for direct intracellular
delivery of
macromolecules comprising MDDT or fragments thereof. For example, liposome
preparations
containing a cell-impermeable macromolecule may promote cell fusion and
intracellular delivery of
the macromolecule. Alternatively, MDDT or a fragment thereof may be joined to
a short cationic N-
terminal portion from the HIV Tat-1 protein. Fusion proteins thus generated
have been found to
transduce into the cells of all tissues, including the brain, in a mouse model
system (Schwarze, S.R. et
al. (1999) Science 285:1569-1572).
For any compound, the therapeutically effective dose can be estimated
initially either in cell
culture assays, e.g., of neoplastic cells, or in animal models such as mice,
rats, rabbits, dogs,
monkeys, or pigs. An animal model may also be used to determine the
appropriate concentration
range and route of administration. Such information can then be used to
determine useful doses and
routes for administration in humans.
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A therapeutically effective dose refers to that amount of active ingredient,
for example
MDDT or fragments thereof, antibodies of MDDT, and agonists, antagonists or
inhibitors of MDDT,
which ameliorates the symptoms or condition. Therapeutic efficacy and toxicity
may be determined
by standard pharmaceutical procedures in cell cultures or with experimental
animals, such as by
calculating the EDSO (the dose therapeutically effective in 50% of the
population) or LDso (the dose
lethal to 50% of the population) statistics. The dose ratio of toxic to
therapeutic effects is the
therapeutic index, which can be expressed as the LDSO/EDSO ratio. Compositions
which exhibit large
therapeutic indices are preferred. The data obtained from cell culture assays
and animal studies are
used to formulate a range of dosage for human use. The dosage contained in
such compositions is
preferably within a range of circulating concentrations that includes the EDso
with little or no toxicity.
The dosage varies within this range depending upon the dosage form employed,
the sensitivity of the
patient, and the route of administration.
The exact dosage will be determined by the practitioner, in light of factors
related to the
subject requiring treatment. Dosage and administration are adjusted to provide
sufficient levels of the
active moiety or to maintain the desired effect. Factors which may be taken
into account include the
severity of the disease state, the general health of the subject, the age,
weight, and gender of the
subject, time and frequency of administration, drug combination(s), reaction
sensitivities, and
response to therapy. Long-acting compositions may be administered every 3 to 4
days, every week,
or biweekly depending on the half-life and clearance rate of the particular
formulation.
Normal dosage amounts may vary from about 0.1 ~g to 100,000 ~cg, up to a total
dose of
about 1 gram, depending upon the route of administration. Guidance as to
particular dosages and
methods of delivery is provided in the literature and generally available to
practitioners in the art.
Those skilled in the art will employ different formulations for nucleotides
than for proteins or their
inhibitors. Similarly, delivery of polynucleotides or polypeptides will be
specific to particular cells,
conditions, locations, etc.
DIAGNOSTICS
In another embodiment, antibodies which specifically bind MDDT may be used for
the
diagnosis of disorders characterized by expression of MDDT, or in assays to
monitor patients being
treated with MDDT or agonists, antagonists, or inhibitors of MDDT. Antibodies
useful for
diagnostic purposes may be prepared in the same manner as described above for
therapeutics.
Diagnostic assays for MDDT include methods which utilize the antibody and a
label to detect MDDT
in human body fluids or in extracts of cells or tissues. The antibodies may be
used with or without
modification, and may be labeled by covalent or non-covalent attachment of a
reporter molecule. A
wide variety of reporter molecules, several of which are described above, are
known in the art and
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may be used.
A variety of protocols for measuring MDDT, including ELISAs, RIAs, and FACS,
are known
in the art and provide a basis for diagnosing altered or abnormal levels of
MDDT expression. Normal
or standard values for MDDT expression are established by combining body
fluids or cell extracts
taken from normal mammalian subjects, for example, human subjects, with
antibodies to MDDT
under conditions suitable for complex formation. The amount of standard
complex formation may be
quantitated by various methods, such as photometric means. Quantities of MDDT
expressed in
subject, control, and disease samples from biopsied tissues are compared with
the standard values.
Deviation between standard and subject values establishes the parameters for
diagnosing disease.
In another embodiment of the invention, polynucleotides encoding MDDT may be
used for
diagnostic purposes. The polynucleotides which may be used include
oligonucleotides,
complementary RNA and DNA molecules, and PNAs. The polynucleotides may be used
to detect
and quantify gene expression in biopsied tissues in which expression of MDDT
may be correlated
with disease. The diagnostic assay may be used to determine absence, presence,
and excess
expression of MDDT, and to monitor regulation of MDDT levels during
therapeutic intervention.
In one aspect, hybridization with PCR probes which are capable of detecting
polynucleotides,
including genomic sequences, encoding MDDT or closely related molecules may be
used to identify
nucleic acid sequences which encode MDDT. The specificity of the probe,
whether it is made from a
highly specific region, e.g., the 5'regulatory region, or from a less specific
region, e.g., a conserved
motif, and the stringency of the hybridization or amplification will determine
whether the probe
identifies only naturally occurring sequences encoding MDDT, allelic variants,
or related sequences.
Probes may also be used for the detection of related sequences, and may have
at least 50%
sequence identity to any of the MDDT encoding sequences. The hybridization
probes of the subject
invention may be DNA or RNA and may be derived from the sequence of SEQ ID
N0:49-96 or from
genomic sequences including promoters, enhancers, and introns of the MDDT
gene.
Means for producing specific hybridization probes for polynucleotides encoding
MDDT
include the cloning of polynucleotides encoding MDDT or MDDT derivatives into
vectors for the
production of mRNA probes. Such vectors are known in the art, are commercially
available, and may
be used to synthesize RNA probes irz vitro by means of the addition of the
appropriate RNA
polymerases and the appropriate labeled nucleotides. Hybridization probes may
be labeled by a
variety of reporter groups, for example, by radionuclides such as 3~P or 355,
or by enzymatic labels,
such as alkaline phosphatase coupled to the probe via avidin/biotin coupling
systems, and the like.
Polynucleotides encoding MDDT may be used for the diagnosis of disorders
associated with
expression of MDDT. Examples of such disorders include, but are not limited
to, a cell proliferative
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disorder such as actinic keratosis, arteriosclerosis, atherosclerosis,
bursitis, cirrhosis, hepatitis, mixed
connective tissue disease (MCTD), myelofibrosis, paroxysmal nocturnal
hemoglobinuria,
polycythemia vera, psoriasis, primary thrombocythemia, and cancers including
adenocarcinoma,
leukemia, lymphoma, melanoma, myeloma, sarcoma, teratocarcinoma, and, in
particular, cancers of
the adrenal gland, bladder, bone, bone marrow, brain, breast, cervix, gall
bladder, ganglia,
gastrointestinal tract, heart, kidney, liver, lung, muscle, ovary, pancreas,
parathyroid, penis, prostate,
salivary glands, skin, spleen, testis, thymus, thyroid, and uterus; an
autoimmune/inflammatory
disorder such as acquired immunodeficiency syndrome (AmS), Addison's disease,
adult respiratory
distress syndrome, allergies, ankylosing spondylitis, amyloidosis, anemia,
asthma, atherosclerosis,
autoimmune hemolytic anemia, autoimmune thyroiditis, autoimmune
polyendocrinopathy-
candidiasis-ectodermal dystrophy (APECED), bronchitis, cholecystitis, contact
dermatitis, Crohn's
disease, atopic dermatitis, dermatomyositis, diabetes mellitus, emphysema,
episodic lymphopenia
with lymphocytotoxins, erythroblastosis fetalis, erythema nodosum, atrophic
gastritis,
glomerulonephritis, Goodpasture's~syndrome, gout, Graves' disease, Hashimoto's
thyroiditis,
hypereosinophilia, irritable bowel syndrome, multiple sclerosis, myasthenia
gravis, myocardial or
pericardial inflammation, osteoarthritis, osteoporosis, pancreatitis,
polymyositis, psoriasis, Reiter's
syndrome, rheumatoid arthritis, scleroderma, Sjogren's syndrome, systemic
anaphylaxis, systemic
lupus erythematosus, systemic sclerosis, thrombocytopenic purpura, ulcerative
colitis, uveitis, Werner
syndrome, complications of cancer, hemodialysis, and extracorporeal
circulation, viral, bacterial,
fungal, parasitic, protozoal, and helminthic infections, and trauma; a
developmental disorder such as
renal tubular acidosis, anemia, Cushing's syndrome, achondroplastic dwarfism,
Duchenne and Becker
muscular dystrophy, epilepsy, gonadal dysgenesis, WAGR syndrome (Wilms' tumor,
aniridia,
genitourinary abnormalities, and mental retardation), Smith-Magenis syndrome,
myelodysplastic
syndrome, hereditary mucoepithelial dysplasia, hereditary keratodermas,
hereditary neuropathies such
as Charcot-Marie-Tooth disease and neurofibromatosis, hypothyroidism,
hydrocephalus, seizure
disorders such as Syndenham s chorea and cerebral palsy, spina bifida,
anencephaly,
craniorachischisis, congenital glaucoma, cataract, and sensorineural hearing
loss; and a neurological
disorder such as epilepsy, ischemic cerebrovascular disease, stroke, cerebral
neoplasms, Alzheimer's
disease, Pick's disease, Huntington's disease, dementia, Parkinson's disease
and other extrapyramidal
disorders, amyotrophic lateral sclerosis and other motor neuron disorders,
progressive neural
muscular atrophy, retinitis pigmentosa, hereditary ataxias, multiple sclerosis
and other demyelinating
diseases, bacterial and viral meningitis, brain abscess, subdural empyema,
epidural abscess,
suppurative intracranial thrombophlebitis, myelitis and radiculitis, viral
central nervous system
disease, prion diseases including kuru, Creutzfeldt-Jakob disease, and
Gerstmann-
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Straussler-Scheinker syndrome, fatal familial insomnia, nutritional and
metabolic diseases of the
nervous system, neurofibromatosis, tuberous sclerosis, cerebelloretinal
hemangioblastomatosis,
encephalotrigeminal syndrome, mental retardation and other developmental
disorders of the central
nervous system including Down syndrome, cerebral palsy, neuroskeletal
disorders, autonomic
nervous system disorders, cranial nerve disorders, spinal cord diseases,
muscular dystrophy and other
neuromuscular disorders, peripheral nervous system disorders, dermatomyositis
and polymyositis,
inherited, metabolic, endocrine, and toxic myopathies, myasthenia gravis,
periodic paralysis, mental
disorders including mood, anxiety, and schizophrenic disorders, seasonal
affective disorder (SAD),
akathesia, amnesia, catatonia, diabetic neuropathy, tardive dyskinesia,
dystonias, paranoid psychoses,
postherpetic neuralgia, Tourette's disorder, progressive supranuclear palsy,
corticobasal degeneration,
and familial frontotemporal dementia. Polynucleotides encoding MDDT may be
used in Southern or
northern analysis, dot blot, or other membrane-based technologies; in PCR
technologies; in dipstick,
pin, and multiformat ELISA-like assays; and in microarrays utilizing fluids or
tissues from patients to
detect altered MDDT expression. Such qualitative or quantitative methods are
well known in the art.
In a particular embodiment, polynucleotides encoding MDDT may be used in
assays that
detect the presence of associated disorders, particularly those mentioned
above. Polynucleotides
complementary to sequences encoding MDDT may be labeled by standard methods
and added to a
fluid or tissue sample from a patient under conditions suitable for the
formation of hybridization
complexes. After a suitable incubation period, the sample is washed and the
signal is quantified and
compared with a standard value. If the amount of signal in the patient sample
is significantly altered
in comparison to a control sample then the presence of altered levels of
polynucleotides encoding
MDDT in the sample indicates the presence of the associated disorder. Such
assays may also be used
to evaluate the efficacy of a particular therapeutic treatment regimen in
animal studies, in clinical
trials, or to monitor the treatment of an individual patient.
In order to provide a basis for the diagnosis of a disorder associated with
expression of
MDDT, a normal or standard profile for expression is established. This may be
accomplished by
combining body fluids or cell extracts taken from normal subjects, either
animal or human, with a
sequence, or a fragment thereof, encoding MDDT, under conditions suitable for
hybridization or
amplification. Standard hybridization may be quantified by comparing the
values obtained from
normal subjects with values from an experiment in which a known amount of a
substantially purified
polynucleotide is used. Standard values obtained in this manner may be
compared with values
obtained from samples from patients who are symptomatic for a disorder.
Deviation from standard
values is used to establish the presence of a disorder.
Once the presence of a disorder is established and a treatment protocol is
initiated,
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hybridization assays may be repeated on a regular basis to determine if the
level of expression in the
patient begins to approximate that which is observed in the normal subject.
The results obtained from
successive assays may be used to show the efficacy of treatment over a period
ranging from several
days to months.
With respect to cancer, the presence of an abnormal amount of transcript
(either under- or
overexpressed) in biopsied tissue from an individual may indicate a
predisposition for the
development of the disease, or may provide a means for detecting the disease
prior to the appearance
of actual clinical symptoms. A more definitive diagnosis of this type may
allow health professionals
to employ preventative measures or aggressive treatment earlier, thereby
preventing the development
or further progression of the cancer.
Additional diagnostic uses for oligonucleotides designed from the sequences
encoding
MDDT may involve the use of PCR. These oligomers may be chemically
synthesized, generated
enzymatically, or produced in vitro. Oligomers will preferably contain a
fragment of a polynucleotide
encoding MDDT, or a fragment of a polynucleotide complementary to the
polynucleotide encoding
MDDT, and will be employed under optimized conditions for identification of a
specific gene or
condition. Oligomers may also be employed under less stringent conditions for
detection or
quantification of closely related DNA or RNA sequences.
In a particular aspect, oligonucleotide primers derived from polynucleotides
encoding MDDT
may be used to detect single nucleotide polymorphisms (SNPs). SNPs are
substitutions, insertions
and deletions that are a frequent cause of inherited or acquired genetic
disease in humans. Methods
of SNP detection include, but are not limited to, single-stranded conformation
polymorphism (SSCP)
and fluorescent SSCP (fSSCP) methods. In SSCP, oligonucleotide primers derived
from
polynucleotides encoding MDDT are used to amplify DNA using the polymerase
chain reaction
(PCR). The DNA may be derived,~for example, from diseased or normal tissue,
biopsy samples,
bodily fluids, and the like. SNPs in the DNA cause differences in the
secondary and tertiary
structures of PCR products in single-stranded form, and these differences are
detectable using gel
electrophoresis in non-denaturing gels. In fSCCP, the oligonucleotide primers
are fluorescently
labeled, which allows detection of the amplimers in high-throughput equipment
such as DNA
sequencing machines. Additionally, sequence database analysis methods, termed
in silico SNP
(isSNP), are capable of identifying polymorphisms by comparing the sequence of
individual
overlapping DNA fragments which assemble into a common consensus sequence.
These computer-
based methods filter out sequence variations due to laboratory preparation of
DNA and sequencing
errors using statistical models and automated analyses of DNA sequence
chromatograms. In the
alternative, SNPs may b~ detected and characterized by mass spectrometry
using, for example, the
5s
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high throughput MASSARRAY system (Sequenom, Inc., San Diego CA).
SNPs may be used to study the genetic basis of human disease. For example, at
least 16
common SNPs have been associated with non-insulin-dependent diabetes mellitus.
SNPs are also
useful for examining differences in disease outcomes in monogenic disorders,
such as cystic fibrosis,
sickle cell anemia, or chronic granulomatous disease. For example, variants in
the mannose-binding
lectin, MBL2, have been shown to be correlated with deleterious pulmonary
outcomes in cystic
fibrosis. SNPs also have utility in pharmacogenomics, the identification of
genetic variants that
influence a patient's response to a drug, such as life-threatening toxicity.
For example, a variation in
N-acetyl transferase is associated with a high incidence of peripheral
neuropathy in response to the
anti-tuberculosis drug isoniazid, while a variation in the core promoter of
the ALOXS gene results in
diminished clinical response to treatment with an anti-asthma drug that
targets the 5-lipoxygenase
pathway. Analysis of the distribution of SNPs in different populations is
useful for investigating
genetic drift, mutation, recombination, and selection, as well as for tracing
the origins of populations
and their migrations (Taylor, J.G. et al. (2001) Trends Mol. Med. 7:507-512;
I~wok, P.-Y. and Z. Gu
(1999) Mol. Med. Today 5:538-543; Nowotny, P. et al. (2001) Curr. Opin.
Neurobiol. 11:637-641).
Methods which may also be used to quantify the expression of MDDT include
radiolabeling
or biotinylating nucleotides, coamplification of a control nucleic acid, and
interpolating results from
standard curves (Melby, P.C. et al. (1993) J. Tmmunol. Methods 159:235-244;
Duplaa, C. et al. (1993)
Anal. Biochem. 212:229-236). The speed of quantitation of multiple samples may
be accelerated by
running the assay in a high-throughput format where the oligomer or
polynucleotide of interest is
presented in various dilutions and a spectrophotometric or colorimetric
response gives rapid
quantitation.
In further embodiments, oligonucleotides or longer fragments derived from any
of the
polynucleotides described herein may be used as elements on a microarray. The
microarray can be
used in transcript imaging techniques which monitor the relative expression
levels of large numbers
of genes simultaneously as described below. The microarray may also be used to
identify genetic
variants, mutations, and polymorphisms. This information may be used to
determine gene function,
to understand the genetic basis of a disorder, to diagnose a disorder, to
monitor
progression/regression of disease as a function of gene expression, and to
develop and monitor the
activities of therapeutic agents in the treatment of disease. In particular,
this information may be used
to develop a pharmacogenomic profile of a patient in order to select the most
appropriate and
effective treatment regimen for that patient. For example, therapeutic agents
which are highly
effective and display the fewest side effects may be selected for a patient
based on his/her
pharmacogenomic profile.
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In another embodiment, MDDT, fragments of MDDT, or antibodies specific for
MDDT may
be used as elements on a microarray. The microarray may be used to monitor or
measure protein-
protein interactions, drug-target interactions, and gene expression profiles,
as described above.
A particular embodiment relates to the use of the polynucleotides of the
present invention to
generate a transcript image of a tissue or cell type. A transcript image
represents the global pattern of
gene expression by a particular tissue or cell type. Global gene expression
patterns are analyzed by
quantifying the number of expressed genes and their relative abundance under
given conditions and at
a given time (Seilhamer et al., "Comparative Gene Transcript Analysis," U.S.
Patent No. 5,840,484;
hereby expressly incorporated by reference herein). Thus a transcript image
may be generated by
hybridizing the polynucleotides of the present invention or their complements
to the totality of
transcripts or reverse transcripts of a particular tissue or cell type. In one
embodiment, the
hybridization takes place in high-throughput format, wherein the
polynucleotides of the present
invention or their complements comprise a subset of a plurality of elements on
a microarray. The
resultant transcript image would provide a profile of gene activity.
Transcript images may be generated using transcripts isolated from tissues,
cell lines,
biopsies, or other biological samples. The transcript image may thus reflect
gene expression ira vivo,
as in the case of a tissue or biopsy sample, or in vitro, as in the case of a
cell line.
Transcript images which profile the expression of the polynucleotides of the
present
invention may also be used in conjunction with in vitro model systems and
preclinical evaluation of
pharmaceuticals, as well as toxicological testing of industrial and naturally-
occurring environmental
compounds. All compounds induce characteristic gene expression patterns,
frequently termed
molecular fingerprints or toxicant signatures, which are indicative of
mechanisms of action and
toxicity (Nuwaysir, E.F. et al. (1999) Mol. Carcinog. 24:153-159; Steiner, S.
and N.L. Anderson
(2000) Toxicol. Lett. 112-113:467-471). If a test compound has a signature
similar to that of a
compound with known toxicity, it is likely to share those toxic properties.
These fingerprints or
signatures are most useful and refined when they contain expression
information from a large number
of genes and gene families. Ideally, a genome-wide measurement of expression
provides the highest
quality signature. Even genes whose expression is not altered by any tested
compounds are important
as well, as the levels of expression of these genes are used to normalize the
rest of the expression
data. The normalization procedure is useful for comparison of expression data
after treatment with
different compounds. While the assignment of gene function to elements of a
toxicant signature aids
in interpretation of toxicity mechanisms, knowledge of gene function is not
necessary for the
statistical matching of signatures which leads to prediction of toxicity (see,
for example, Press
Release 00-02 from the National Institute of Environmental Health Sciences,
released February 29,
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2000, available at http:/lwww.niehs.nih.gov/oclnewsltoxchip.htm). Therefore,
it is important and
desirable in toxicological screening using toxicant signatures to include all
expressed gene sequences.
In an embodiment, the toxicity of a test compound can be assessed by treating
a biological
sample containing nucleic acids with the test compound. Nucleic acids that are
expressed in the
treated biological sample are hybridized with one or more probes specific to
the polynucleotides of
the present invention, so that transcript levels corresponding to the
polynucleotides of the present
invention may be quantified. The transcript levels in the treated biological
sample are compared with
levels in an untreated biological sample. Differences in the transcript levels
between the two samples
are indicative of a toxic response caused by the test compound in the treated
sample.
Another embodiment relates to the use of the polypeptides disclosed herein to
analyze the
proteome of a tissue or cell type. The term proteome refers to the global
pattern of protein expression
in a particular tissue or cell type. Each protein component of a proteome can
be subjected
individually to further analysis. Proteome expression patterns, or profiles,
are analyzed by
quantifying the number of expressed proteins and their relative abundance
under given conditions and
at a given time. A profile of a cell's proteome may thus be generated by
separating and analyzing the
polypeptides of a particular tissue or cell type. In one embodiment, the
separation is achieved using
two-dimensional gel electrophoresis, in which proteins from a sample are
separated by isoelectric
focusing in the first dimension, and then according to molecular weight by
sodium dodecyl sulfate
slab gel electrophoresis in the second dimension (Steiner and Anderson,
suyra). The proteins are
visualized in the gel as discrete and uniquely positioned spots, typically by
staining the gel with an
agent such as Coomassie Blue or silver or fluorescent stains. The optical
density of each protein spot
is generally proportional to the level of the protein in the sample. The
optical densities of
equivalently positioned protein spots from different samples, for example,
from biological samples
either treated or untreated with a test compound or therapeutic agent, are
compared to identify any
changes in protein spot density related to the treatment. The proteins in the
spots are partially
sequenced using, for example, standard methods employing chemical or enzymatic
cleavage followed
by mass spectrometry. The identity of the protein in a spot may be determined
by comparing its
partial sequence, preferably of at least 5 contiguous amino acid residues, to
the polypeptide sequences
of interest. In some cases, further sequence data may be obtained for
definitive protein identification.
A proteomic profile may also be generated using antibodies specific for MDDT
to quantify
the levels of MDDT expression. In one embodiment, the antibodies are used as
elements on a
microarray, and protein expression levels are quantified by exposing the
microarray to the sample and
detecting the levels of protein bound to each array element (Luehing, A. et
al. (1999) Anal. Biochem.
270:103-111; Mendoze, L.G. et al. (1999) Biotechniques 27:778-788). Detection
may be performed
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by a variety of methods known in the art, for example, by reacting the
proteins in the sample with a
thiol- or amino-reactive fluorescent compound and detecting the amount of
fluorescence bound at
each array element.
Toxicant signatures at the proteome level are also useful for toxicological
screening, and
should be analyzed in parallel with toxicant signatures at the transcript
level. There is a poor
correlation between transcript and protein abundances for some proteins in
some tissues (Anderson,
N.L. and J. Seilhamer (1997) Electrophoresis 18:533-537), so proteome toxicant
signatures may be
useful in the analysis of compounds which do not significantly affect the
transcript image, but which
alter the proteomic profile. In addition, the analysis of transcripts in body
fluids is difficult, due to
rapid degradation of mRNA, so proteomic profiling may be more reliable and
informative in such
cases.
In another embodiment, the toxicity of a test compound is assessed by treating
a biological
sample containing proteins with the test compound. Proteins that are expressed
in the treated
biological sample are separated so that the amount of each protein can be
quantified. The amount of
15. each protein is compared to the amount of the corresponding protein in an
untreated biological
sample. A difference in the amount of protein between the two samples is
indicative of a toxic
response to the test compound in the treated sample. Individual proteins are
identified by sequencing
the amino acid residues of the individual proteins and comparing these partial
sequences to the
polypeptides of the present invention.
In another embodiment, the toxicity of a test compound is assessed by treating
a biological
sample containing proteins with the test compound. Proteins from the
biological sample are
incubated with antibodies specific to the polypeptides of the present
invention. The amount of
protein recognized by the antibodies is quantified. The amount of protein in
the treated biological
sample is compared with the amount in an untreated biological sample. A
difference in the amount of
protein between the two samples is indicative of a toxic response to the test
compound in the treated
sample.
Microarrays may be prepared, used, and analyzed using methods known in the art
(Brennan,
T.M. et al. (1995) U.S. Patent No. 5,474,796; Schena, M. et al. (1996) Proc.
Natl. Acad. Sci. USA
93:10614-10619; Baldeschweiler et al. (1995) PCT application W095/251116;
Shalom D. et al.
(1995) PCT application W095/35505; Heller, R.A. et al. (1997) Proc. Natl.
Acad. Sci. USA 94:2150-
2155; Heller, M.J. et al. (1997) U.S. Patent No. 5,605,662). Various types of
microarrays are well
known and thoroughly described in Schena, M., ed. (1999; DNA Microarrays: A
Practical Apt~roach,
Oxford University Press, London).
In another embodiment of the invention, nucleic acid sequences encoding MDDT
may be
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used to generate hybridization probes useful in mapping the naturally
occurring genomic sequence.
Either coding or noncoding sequences may be used, and in some instances,
noncoding sequences may
be preferable over coding sequences. For example, conservation of a coding
sequence among
members of a mufti-gene family may potentially cause undesired cross
hybridization during
chromosomal mapping. The sequences may be mapped to a particular chromosome,
to a specific
region of a chromosome, or to artificial chromosome constructions, e.g., human
artificial
chromosomes (HACs), yeast artificial chromosomes (YACs), bacterial artificial
chromosomes
(BACs), bacterial P1 constructions, or single chromosome cDNA libraries
(Harrington, J.J. et al.
(1997) Nat. Genet. 15:345-355; Price, C.M. (1993) Blood Rev. 7:127-134; Trask,
B.J. (1991) Trends
Genet. 7:149-154). Once mapped, the nucleic acid sequences may be used to
develop genetic linkage
maps, for example, which correlate the inheritance of a disease state with the
inheritance of a
particular chromosome region or restriction fragment length polymorphism
(RFLP) (Lander, E.S. and
D. Botstein (1986) Proc. Natl. Acad. Sci. USA 83:7353-7357).
Fluorescent irz situ hybridization (FISH) may be correlated with other
physical and genetic
map data (Heinz-Ulrich, et al. (1995) in Meyers, supra, pp. 965-968). Examples
of genetic map data
can be found in various scientific journals or at the Online Mendelian
Inheritance in Man (OMIM)
World Wide Web site. Correlation between the location of the gene encoding
MDDT on a physical
map and a specific disorder, or a predisposition to a specific disorder, may
help define the region of
DNA associated with that disorder and thus may further positional cloning
efforts.
Izz situ hybridization of chromosomal preparations and physical mapping
techniques, such as
linkage analysis using established chromosomal markers, may be used for
extending genetic maps.
Often the placement of a gene on the chromosome of another mammalian species,
such as mouse,
may reveal associated markers even if the exact chromosomal locus is not
known. This information
is valuable to investigators searching for disease genes using positional
cloning or other gene
discovery techniques. Once the gene or genes responsible for a disease or
syndrome have been
crudely localized by genetic linkage to a particular genomic region, e.g.,
ataxia-telangiectasia to
11q22-23, any sequences mapping to that area may represent associated or
regulatory genes for
further investigation (Gatti, R.A. et al. (1988) Nature 336:577-580). The
nucleotide sequence of the
instant invention may also be used to detect differences in the chromosomal
location due to
translocation, inversion, etc., among normal, carrier, or affected
individuals.
In another embodiment of the invention, MDDT, its catalytic or immunogenic
fragments, or
oligopeptides thereof can be used for screening libraries of compounds iri any
of a variety of drug
screening techniques. The fragment employed in such screening may be free in
solution, affixed to a
solid support, borne on a cell surface, or located intracellularly. The
formation of binding complexes
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between MDDT and the agent being tested may be measured.
Another technique for drug screening provides for high throughput screening of
compounds
having suitable binding affinity to the protein of interest (Geysen, et al.
(1984) PCT application
W084103564). In this method, large numbers of different small test compounds
are synthesized on a
solid substrate. The test compounds axe reacted with MDDT, or fragments
thereof, and washed.
Bound MDDT is then detected by methods well known in the art. Purified MDDT
can also be coated
directly onto plates for use in the aforementioned drug screening techniques.
Alternatively,
non-neutralizing antibodies can be used to capture the peptide and immobilize
it on a solid support.
In another embodiment, one may use competitive drug screening assays in which
neutralizing
antibodies capable of binding MDDT specifically compete with a test compound
for binding MDDT.
In this manner, antibodies can be used to detect the presence of any peptide
which shares one or more
antigenic determinants with MDDT.
In additional embodiments, the nucleotide sequences which encode MDDT may be
used in
any molecular biology techniques that have yet to be developed, provided the
new techniques rely on
properties of nucleotide sequences that are currently known, including, but
not limited to, such
properties as the triplet genetic code and specific base pair interactions.
Without further elaboration, it is believed that one skilled in the art can,
using the preceding
description, utilize the present invention to its fullest extent. The
following embodiments are,
therefore, to be construed as merely illustrative, and not !imitative of the
remainder of the disclosure
in any way whatsoever.
The disclosures of all patents, applications, and publications mentioned above
and below,
including U.S. Ser. No. 60/328,944, U.S. Ser. No. 60/332,430, U.S. Ser. No.
60/343,880, U.S. Ser.
No. 60!345,143, and U.S. Ser. No. 60/345,384, are hereby expressly
incorporated by reference.
EXAMPLES
I. Construction of cDNA Libraries
Incyte cDNAs were derived from cDNA libraries described in the L1FESEQ GOLD
database
(Incyte Genomics, Palo Alto CA). Some tissues were homogenized and lysed in
guanidinium
isothiocyanate, while others were homogenized and lysed in phenol or in a
suitable mixture of
denaturants, such as TRIZOL (Invitrogen), a monophasic solution of phenol and
guanidine
isothiocyanate. The resulting lysates were centrifuged over CsCl cushions or
extracted with
chloroform. RNA was precipitated from the lysates with either isopropanol or
sodium acetate and
ethanol, or by other routine methods.
Phenol extraction and precipitation of RNA were repeated as necessary to
increase RNA
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purity. In some cases, RNA was treated with DNase. For most libraries,
poly(A)+ RNA was isolated
using oligo d(T)-coupled paramagnetic particles (Promega), OLIGOTEX latex
particles (QIAGEN,
Chatsworth CA), or an OLIGOTEX mRNA purification kit (QIAGEN). Alternatively,
RNA was
isolated directly from tissue lysates using other RNA isolation kits, e.g.,
the POLY(A)PURE mRNA
purification kit (Ambion, Austin TX).
In some cases, Stratagene was provided with RNA and constructed the
corresponding cDNA
libraries. Otherwise, cDNA was synthesized and cDNA libraries were constructed
with the UNIZAP
vector system (Stratagene) or SUPERSCRIPT plasmid system (hivitrogen), using
the recommended
procedures or similar methods known in the art (Ausubel et al., supra, ch. 5).
Reverse transcription
was initiated using oligo d(T) or random primers. Synthetic oligonucleotide
adapters were ligated to
double stranded cDNA, and the cDNA was digested with the appropriate
restriction enzyme or
enzymes. For most libraries, the cDNA was size-selected (300-1000 bp) using
SEPHACRYL S 1000,
SEPHAROSE CL2B, or SEPHAROSE CL4B column chromatography (Amersham Biosciences)
or
preparative agarose gel electrophoresis. cDNAs were ligated into compatible
restriction enzyme sites
of the polylinker of a suitable plasmid, e.g., PBLUESCRIPT plasmid
(Stratagene), PSPORT1 plasmid
(Invitrogen, Carlsbad CA), PCDNA2.1 plasmid (Invitrogen), PBK-CMV plasmid
(Stratagene), PCR2-
TOPOTA plasmid (Invitrogen), PCMV-ICIS plasmid (Stratagene), pIGEN (fiicyte
Genomics, Palo
Alto CA), pRARE (Incyte Genomics), or pINCY (Incyte Genomics), or derivatives
thereof.
Recombinant plasmids were transformed into competent E. coli cells including
XL1-Blue, XL1-
BIueMRF, or SOLR from Stratagene or DHSa, DHlOB, or ElectroMAX DH10B from
Invitrogen.
II. Isolation of cDNA Clones
Plasmids obtained as described in Example I were recovered from host cells by
iu vivo
excision using the UNIZAP vector system (Stratagene) or by cell lysis.
Plasmids were purified using
at least one of the following: a Magic or WIZARD Minipreps DNA purification
system (Promega);
an AGTC Miniprep purification kit (Edge Biosystems, Gaithersburg MD); and
QIAWELL 8 Plasmid,
QIAWELL 8 Plus Plasmid, QIAWELL 8 Ultra Plasmid purification systems or the
R.E.A.L. PREP 96
plasmid purification kit from QIAGEN. Following precipitation, plasmids were
resuspended in 0.1
ml of distilled water and stored, with or without lyophilization, at
4°C.
Alternatively, plasmid DNA was amplified from host cell lysates using direct
link PCR in a
high-throughput format (Rao, V.B. (1994) Anal. Biochem. 216:1-14). Host cell
lysis and thermal
cycling steps were carried out in a single reaction mixture. Samples were
processed and stored in
384-well plates, and the concentration of amplified plasmid DNA was quantified
fluorometrically
using PICOGREEN dye (Molecular Probes, Eugene OR) and a FLUOROSKAN II
fluorescence
scanner (Labsystems Oy, Helsinki, Finland).
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III. Sequencing and Analysis
Incyte cDNA recovered in plasmids as described in Example II were sequenced as
follows.
Sequencing reactions were processed using standard methods or high-throughput
instrumentation
such as the ABI CATALYST 800 (Applied Biosystems) thermal cycler or the PTC-
200 thermal
cycler (MJ Research) in conjunction with the HYDRA microdispenser (Robbins
Scientific) or the
MICROLAB 2200 (Hamilton) liquid transfer system. cDNA sequencing reactions
were prepared
using reagents provided by Amersham Biosciences or supplied in ABI sequencing
kits such as the
ABI PRISM BIGDYE Terminator cycle sequencing ready reaction kit (Applied
Biosystems).
Electrophoretic separation of cDNA sequencing reactions and detection of
labeled polynucleotides
were carried out using the MEGABACE 1000 DNA sequencing system (Amersham
Biosciences); the
ABI PRISM 373 or 377 sequencing system (Applied Biosystems) in conjunction
with standard ABI
protocols and base calling software; or other sequence analysis systems known
in the art. Reading
frames within the cDNA sequences were identified using standard methods
(Ausubel et al., supra, ch.
7). Some of the cDNA sequences were selected for extension using the
techniques disclosed in
Example VIII.
The polynucleotide sequences derived from Incyte cDNAs were validated by
removing
vector, linker, and poly(A) sequences and by masking ambiguous bases, using
algorithms and
programs based on BLAST, dynamic programming, and dinucleotide nearest
neighbor analysis. The
Incyte cDNA sequences or translations thereof were then queried against a
selection of public
databases such as the GenBank primate, rodent, mammalian, vertebrate, and
eukaryote databases, and
BLOCKS, PRINTS, DOMO, PRODOM; PROTEOME databases with sequences from Hofno
sapieras,
Rattus Jzorvegicus, Mus rnusczzlus, Caenorhabditis elegazzs, Sacclzaroszzyces
cer-evisiae,
Sclzizosacclzaromyces pombe, and Cazzdida albicans (Incyte Genomics, Palo Alto
CA); hidden
Markov model (HMM)-based protein family databases such as PFAM, INCY, and
TIGRFAM (Haft,
D.H. et al. (2001) Nucleic Acids Res. 29:41-43); and HMM-based protein domain
databases such as
SMART (Schultz, J. et al. (1998) Proc. Natl. Acad. Sci. USA 95:5857-5864;
Letunic, I. et al. (2002)
Nucleic Acids Res. 30:242-244).' (HMM is a probabilistic approach which
analyzes consensus
primary structures of gene families; see, for example, Eddy, S.R. (1996) Curr.
Opin. Struct. Biol.
6:361-365.) The queries were performed using programs based on BLAST, FASTA,
BLIMPS, and
HMMER. The Incyte cDNA sequences were assembled to produce full length
polynucleotide
sequences. Alternatively, GenBank cDNAs, GenBank ESTs, stitched sequences,
stretched sequences,
or Genscan-predicted coding sequences (see Examples IV and V) were used to
extend Incyte cDNA
assemblages to full length. Assembly was performed using programs based on
Phred, Phrap, and
Consed, and cDNA assemblages were screened for open reading frames using
programs based on
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GeneMark, BLAST, and FASTA. The full length polynucleotide sequences were
translated to derive
the corresponding full length polypeptide sequences. Alternatively, a
polypeptide may begin at any
of the methionine residues of the full length translated polypeptide. Full
length polypeptide
sequences were subsequently analyzed by querying against databases such as the
GenBank protein
databases (genpept), SwissProt, the PROTEOME databases, BLOCKS, PRINTS, DOMO,
PRODOM,
Prosite, hidden Markov model (HMM)-based protein family databases such as
PFAM, INCY, and
TIGRFAM; and HMM-based protein domain databases such as SMART. Full length
polynucleotide
sequences are also analyzed using MACDNASIS PRO software (MiraiBio, Alameda
CA) and
LASERGENE software (DNASTAR). Polynucleotide and polypeptide sequence
alignments are
generated using default parameters specified by the CLUSTAL algorithm as
incorporated into the
MEGALIGN multisequence alignment program (DNASTAR), which also calculates the
percent
identity between aligned sequences.
Table 7 summarizes the tools, programs, and algorithms used for the analysis
and assembly of
Incyte cDNA and full length sequences and provides applicable descriptions,
references, and
threshold parameters. The first column of Table 7 shows the tools, programs,
and algorithms used,
the second column provides brief descriptions thereof, the third column
presents appropriate
references, all of which are incorporated by reference herein in their
entirety, and the fourth column
presents, where applicable, the scores, probability values, and other
parameters used to evaluate the
strength of a match between two sequences (the higher the score or the lower
the probability value,
the greater the identity between two sequences).
The programs described above for the assembly and analysis of full length
polynucleotide
and polypeptide sequences were also used to identify polynucleotide sequence
fragments from SEQ
ID NO:49-96. Fragments from about 20 to about 4000 nucleotides which are
useful in hybridization
and amplification technologies are described in Table 4, column 2.
IV. Identification and Editing of Coding Sequences from Genomic DNA
Putative molecules for disease detection and treatment were initially
identified by running the
Genscan gene identification program against public genomic sequence databases
(e.g., gbpri and
gbhtg). Genscan is a general-purpose gene identification program which
analyzes genomic DNA
sequences from a variety of organisms (Barge, C. and S. Karlin (1997) J. Mol.
Biol. 268:78-94;
Barge, C. and S. Karlin (1998) Curr. Opin. Struct. Biol. 8:346-354). The
program concatenates
predicted exons to form an assembled cDNA sequence extending from a methionine
to a stop codon.
The output of Genscan is a FASTA database of polynucleotide and polypeptide
sequences. The
maximum range of sequence for Genscan to analyze at once was set to 30 kb. To
determine which of
these Genscan predicted cDNA sequences encode molecules for disease detection
and treatment, the
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encoded polypeptides were analyzed by querying against PFAM models for
molecules for disease
detection and treatment. Potential molecules for disease detection and
treatment were also identified
by homology to Incyte cDNA sequences that had been annotated as molecules for
disease detection
and treatment. These selected Genscan-predicted sequences were then compared
by BLAST analysis
to the genpept and gbpri public databases. Where necessary, the Genscan-
predicted sequences were
then edited by comparison to the top BLAST hit from genpept to correct errors
in the sequence
predicted by Genscan, such as extra or omitted exons. BLAST analysis was also
used to find any
Incyte cDNA or public cDNA coverage of the Genscan-predicted sequences, thus
providing evidence
for transcription. When Incyte cDNA coverage was available, this information
was used to correct or
confirm the Genscan predicted sequence. Full length polynucleotide sequences
were obtained by
assembling Genscan-predicted coding sequences with Incyte cDNA sequences
andlor public cDNA
sequences using the assembly process described in Example III. Alternatively,
full length
polynucleotide sequences were derived entirely from edited or unedited Genscan-
predicted coding
sequences.
V. Assembly of Genomic Sequence Data with cDNA Sequence Data
"Stitched" Sequences
Partial cDNA sequences were extended with exons predicted by the Genscan gene
identification program described in Example IV. Partial cDNAs assembled as
described in Example
III were mapped to genomic DNA and parsed into clusters containing related
cDNAs and Genscan
exon predictions from one or more genomic sequences. Each cluster was analyzed
using an algorithm
based on graph theory and dynamic programming to integrate cDNA and genomic
information,
generating possible splice variants that were subsequently confirmed, edited,
or extended to create a
full length sequence. Sequence intervals in which the entire length of the
interval was present on
more than one sequence in the cluster were identified, and intervals thus
identified were considered to
be equivalent by transitivity. For example, if an interval was present on a
cDNA and two genomic
sequences, then all three intervals were considered to be equivalent. This
process allows unrelated
but consecutive genomic sequences to be brought together, bridged by cDNA
sequence. Intervals
thus identified were then "stitched" together by the stitching algorithm in
the order that they appear
along their parent sequences to generate the longest possible sequence, as
well as sequence variants.
Linkages between intervals which proceed along one type of parent sequence
(cDNA to cDNA or
genomic sequence to genomic sequence) were given preference over linkages
which change parent
type (cDNA to genomic sequence). The resultant stitched sequences were
translated and compared
by BLAST analysis to the genpept and gbpri public databases. Incorrect exons
predicted by Genscan
were corrected by comparison to the top BLAST hit from genpept. Sequences were
further extended
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with additional cDNA sequences, or by inspection of genomic DNA, when
necessary.
"Stretched" Sequences
Partial DNA sequences were extended to full length with an algorithm based on
BLAST
analysis. First, partial cDNAs assembled as described in Example III were
queried against public
databases such as the GenBank primate, rodent, mammalian, vertebrate, and
eukaryote databases
using the BLAST program. The nearest GenBank protein homolog was then compared
by BLAST
analysis to either Incyte cDNA sequences or GenScan exon predicted sequences
described in
Example 1V. A chimeric protein was generated by using the resultant high-
scoring segment pairs
(HSPs) to map the translated sequences onto the GenBank protein homolog.
Insertions or deletions
may occur in the chimeric protein with respect to the original GenBank protein
homolog. The
GenBank protein homolog, the chimeric protein, or both were used as probes to
search for
homologous genomic sequences from the public human genome databases. Partial
DNA sequences
were therefore "stretched" or extended by the addition of homologous genomic
sequences. The
resultant stretched sequences were examined to determine whether it contained
a complete gene.
VI. Chromosomal Mapping of MDDT Encoding Polynucleotides
The sequences which were used to assemble SEQ ID N0:49-96 were compared with
sequences from the Incyte LIFESEQ database and public domain databases using
BLAST and other
implementations of the Smith-Waterman algorithm. Sequences from these
databases that matched
SEQ ID NO:49-96 were assembled into clusters of contiguous and overlapping
sequences using
assembly algorithms such as Phrap (Table 7). Radiation hybrid and genetic
mapping data available
from public resources such as the Stanford Human Genome Center (SHGC),
Whitehead Institute for
Genome Research (WIGR), and Genethon were used to determine if any of the
clustered sequences
had been previously mapped. Inclusion of a mapped sequence in a cluster
resulted in the assignment
of all sequences of that cluster, including its particular SEQ ID NO:, to that
map location.
Map locations are represented by ranges, or intervals, of human chromosomes.
The map
position of an interval, in centiMorgans, is measured relative to the terminus
of the chromosome's p-
arm. (The centiMorgan (cM) is a unit of measurement based on recombination
frequencies between
chromosomal markers. On average, 1 cM is roughly equivalent to 1 megabase (Mb)
of DNA in
humans, although this can vary widely due to hot and cold spots of
recombination.) The cM
distances are based on genetic markers mapped by Genethon which provide
boundaries for radiation
hybrid markers whose sequences were included in each of the clusters. Human
genome maps and
other resources available to the public, such as the NCBI "GeneMap'99" World
Wide Web site
(http://www.ncbi.nlm.nih.gov/genemap/), can be employed to determine if
previously identified
disease genes map within or in proximity to the intervals indicated above.
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VII. Analysis of Polynucleotide Expression
Northern analysis is a laboratory technique used to detect the presence of a
transcript of a
gene and involves the hybridization of a labeled nucleotide sequence to a
membrane on which RNAs
from a particular cell type or tissue have been bound (Sambrook and Russell,
supra, ch. 7; Ausubel et
al., supra, ch. 4).
Analogous computer techniques applying BLAST were used to search for identical
or related
molecules in databases such as GenBank or LIFESEQ (Incyte Genomics). This
analysis is much
faster than multiple membrane-based hybridizations. In addition, the
sensitivity of the computer
search can be modified to determine whether any particular match is
categorized as exact or similar.
The basis of the search is the product score, which is defined as:
BLAST Score x Percent Identity
5 x minimum {length(Seq. 1), length(Seq. 2)}
The product score takes into account both the degree of similarity between two
sequences and the
length of the sequence match. The product score is a normalized value between
0 and 100, and is
calculated as follows: the BLAST score is multiplied by the percent nucleotide
identity and the
product is divided by (5 times the length of the shorter of the two
sequences). The BLAST score is
calculated by assigning a score of +5 for every base that matches in a high-
scoring segment pair
(HSP), and -4 for every mismatch. Two sequences may share more than one HSP
(separated by
gaps). If there is more than one HSP, then the pair with the highest BLAST
score is used to calculate
the product score. The product score represents a balance between fractional
overlap and quality in a
BLAST alignment. For example, a product score of 100 is produced only for 100%
identity over the
entire length of the shorter of the two sequences being compared. A product
score of 70 is produced
either by 100% identity and 70% overlap at one end, or by 88% identity and
100% overlap at the
other. A product score of 50 is produced either by 100% identity and 50%
overlap at one end, or 79%
identity and 100% overlap.
Alternatively, polynucleotides encoding MDDT are analyzed with respect to the
tissue
sources from which they were derived. For example, some full length sequences
are assembled, at
least in part, with overlapping Incyte cDNA sequences (see Example III). Each
cDNA sequence is
derived from a cDNA library constructed from a human tissue. Each human tissue
is classified into
one of the following organ/tissue categories: cardiovascular system;
connective tissue; digestive
system; embryonic structures; endocrine system; exocrine glands; genitalia,
female; genitalia, male;
germ cells; heroic and immune system; liver; musculoskeletal system; nervous
system; pancreas;
respiratory system; sense organs; skin; stomatognathic system;
unclassified/mixed; or urinary tract.
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The number of libraries in each category is counted and divided by the total
number of libraries
across all categories. Similarly, each human tissue is classified into one of
the following
disease/condition categories: cancer, cell line, developmental, inflammation,
neurological, trauma,
cardiovascular, pooled, and other, and the number of libraries in each
category is counted and divided
by the total number of libraries across all categories. The resulting
percentages reflect the tissue- and
disease-specific expression of cDNA encoding MDDT. cDNA sequences and cDNA
library/tissue
information are found in the LIFESEQ GOLD database (Incyte Genomics, Palo Alto
CA).
VIII. Extension of MDDT Encoding Polynucleotides
Full length polynucleotides are produced by extension of an appropriate
fragment of the full
length molecule using oligonucleotide primers designed from this fragment. One
primer was
synthesized to initiate 5' extension of the known fragment, and the other
primer was synthesized to
initiate 3' extension of the known fragment. The initial primers were designed
using OLIGO 4.06
software (National Biosciences), or another appropriate program, to be about
22 to 30 nucleotides in
length, to have a GC content of about 50% or more, and to anneal to the target
sequence at
temperatures of about 68°C to about 72°C. Any stretch of
nucleotides which would result in hairpin
structures and primer-primer dimerizations was avoided.
Selected human cDNA libraries were used to extend the sequence. If more than
one
extension was necessary or desired, additional or nested sets of primers were
designed.
High fidelity amplification was obtained by PCR using methods well known in
the art. PCR
was performed in 96-well plates using the PTC-200 thermal cycler (MJ Research,
Inc.). The reaction
mix contained DNA template, 200 nmol of each primer, reaction buffer
containing Mgz+, (NH4)zS04,
and 2-mercaptoethanol, Taq DNA polymerase (Amersham Biosciences), ELONGASE
enzyme
(Invitrogen), and Pfu DNA polymerase (Stratagene), with the following
parameters for primer pair
PCI A and PCI B : Step 1: 94 ° C, 3 min; Step 2: 94 ° C, 15 sec;
Step 3: 60 ° C, 1 min; Step 4: 68 ° C, 2
min; Step 5: Steps 2, 3, and 4 repeated 20 times; Step 6: 68°C, 5 min;
Step 7: storage at 4°C. In the
alternative, the parameters for primer pair T7 and SK+ were as follows: Step
1: 94°C, 3 min; Step 2:
94°C, 15 sec; Step 3: 57°C, 1 min; Step 4: 68°C, 2 min;
Step 5: Steps 2, 3, and 4 repeated 20 times;
Step 6: 68°C, 5 min; Step 7: storage at 4°C.
The concentration of DNA in each well was determined by dispensing 100 ~.l
PICOGREEN
quantitation reagent (0.25% (v/v) PICOGREEN; Molecular Probes, Eugene OR)
dissolved in 1X TE
and 0.5 ~.1 of undiluted PCR product into each well of an opaque fluorimeter
plate (Corning Costar,
Acton MA), allowing the DNA to bind to the reagent. The plate was scanned in a
Fluoroskan II
(Labsystems Oy, Helsinki, Finland) to measure the fluorescence of the sample
and to quantify the
concentration of DNA. A 5 ~l to 10 ,u1 aliquot of the reaction mixture was
analyzed by
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electrophoresis on a 1 % agarose gel to determine which reactions were
successful in extending the
sequence.
The extended nucleotides were desalted and concentrated, transferred to 384-
well plates,
digested with CviJI cholera virus endonuclease (Molecular Biology Research,
Madison WI), and
sonicated or sheared prior to religation into pUC 18 vector (Amersham
Biosciences). For shotgun
sequencing, the digested nucleotides were separated on low concentration (0.6
to 0.8%) agarose gels,
fragments were excised, and agar digested with Agar ACE (Promega). Extended
clones were
religated using T4 ligase (New England Biolabs, Beverly MA) into pUC 18 vector
(Amersham
Biosciences), treated with Pfu DNA polymerase (Stratagene) to fill-in
restriction site overhangs, and
transfected into competent E. coli cells. Transformed cells were selected on
antibiotic-containing
media, and individual colonies were picked and cultured overnight at
37°C in 384-well plates in
LB/2x carb liquid media.
The cells were lysed, and DNA was amplified by PCR using Taq DNA polymerase
(Amersham Biosciences) and Pfu DNA polymerase (Stratagene) with the following
parameters: Step
1: ~ 94 ° C, 3 min; Step 2: 94 ° C, 15 sec; Step 3: 60 °
C, 1 min; Step 4: 72 ° C, 2 min; Step 5 : steps 2, 3,
and 4 repeated 29 times; Step 6: 72°C, 5 min; Step 7: storage at
4°C. DNA was quantified by
PICOGREEN reagent (Molecular Probes) as described above. Samples with low DNA
recoveries
were reamplified using the same conditions as described above. Samples were
diluted with 20%
dimethysulfoxide (1:2, v/v), and sequenced using DYENAMIC energy transfer
sequencing primers
and the DYENAMIC DIRECT kit (Amersham Biosciences) or the ABI PRISM BIGDYE
Terminator
cycle sequencing ready reaction kit (Applied Biosystems).
In like manner, full length polynucleotides are verified using the above
procedure or are used
to obtain 5' regulatory sequences using the above procedure along with
oligonucleotides designed for
such extension, and an appropriate genomic library.
IX. Identification of Single Nucleotide Polymorphisms in MDDT Encoding
Polynucleotides
Common DNA sequence variants known as single nucleotide polymorphisms (SNPs)
were
identified in SEQ ID N0:49-96 using the LIF'ESEQ database (Incyte Genomics).
Sequences from the
same gene were clustered together and assembled as described in Example III,
allowing the
identification of all sequence variants in the gene. An algorithm consisting
of a series of filters was
used to distinguish SNPs from other sequence variants. Preliminary filters
removed the majority of
basecall errors by requiring a minimum Phred quality score of 15, and removed
sequence alignment
errors and errors resulting from improper trimming of vector sequences,
chimeras, and splice
variants. An automated procedure of advanced chromosome analysis analysed the
original
chromatogram files in the vicinity of the putative SNP. Clone error filters
used statistically generated
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algorithms to identify errors introduced during laboratory processing, such as
those caused by reverse
transcriptase, polymerase, or somatic mutation. Clustering error filters used
statistically generated
algorithms to identify errors resulting from clustering of close homologs or
pseudogenes, or due to
contamination by non-human sequences. A final set of filters removed
duplicates and SNPs found in
immunoglobulins or T-cell receptors.
Certain SNPs were selected for further characterization by mass spectrometry
using the high
throughput MASSARRAY system (Sequenom, Inc.) to analyze allele frequencies at
the SNP sites in
four different human populations. The Caucasian population comprised 92
individuals (46 male, 46 ~~',°'
female), including 83 from Utah, four French, three Venezualan, and two Amish
individuals. The
African population comprised 194 individuals (97 male, 97 female), all African
Americans. The
Hispanic population comprised 324 individuals (162 male, 162 female), all
Mexican Hispanic. The
Asian population comprised 126 individuals (64 male, 62 female) with a
reported parental breakdown
of 43% Chinese, 31% Japanese, 13% Korean, 5% Vietnamese, and 8% other Asian.
Allele
frequencies were first analyzed in the Caucasian population; in some cases
those SNPs which showed
no allelic variance in this population were not further tested in the other
three populations.
X. Labeling and Use of Individual Hybridization Probes
Hybridization probes derived from SEQ ID N0:49-96 are employed to screen
cDNAs,
genomic DNAs, or mRNAs. Although the labeling of oligonucleotides, consisting
of about 20 base
pairs, is specifically described, essentially the same procedure is used with
larger nucleotide
fragments. Oligonucleotides are designed using state-of-the-art software such
as OLIGO 4.06
software (National Biosciences) and labeled by combining 50 pmol of each
oligomer, 250 ~Ci of
[y-32P~ adenosine triphosphate (Amersham Biosciences), and T4 polynucleotide
kinase (DuPont NEN,
Boston MA). The labeled oligonucleotides are substantially purified using a
SEPHADEX G-25
superfine size exclusion dextran bead column (Amersham Biosciences). An
aliquot containing 10'
counts per minute of the labeled probe is used in a typical membrane-based
hybridization analysis of
human genomic DNA digested with one of the following endonucleases: Ase I, Bgl
II, Eco RI, Pst I,
Xba I, or Pvu II (DuPont NEN).
The DNA from each digest is fractionated on a 0.7% agarose gel and transferred
to nylon
membranes (Nytran Plus, Schleicher & Schuell, Durham NH). Hybridization is
carried out for 16
hours at 40°C. To remove nonspecific signals, blots are sequentially
washed at room temperature
under conditions of up to, for example, 0.1 x saline sodium citrate and 0.5%
sodium dodecyl sulfate.
Hybridization patterns are visualized using autoradiography or an alternative
imaging means and
compared.
XI. Microarrays
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The linkage or synthesis of array elements upon a microarray can be achieved
utilizing
photolithography, piezoelectric printing (ink jet printing; see, e.g.,
Baldeschweiler et al., supra),
mechanical microspotting technologies, and derivatives thereof. The substrate
in each of the
aforementioned technologies should be uniform and solid with a non-porous
surface (Schena, M., ed.
(1999) DNA Microarrays: A Practical Approach, Oxford University Press,
London). Suggested
substrates include silicon, silica, glass slides, glass chips, and silicon
wafers. Alternatively, a
procedure analogous to a dot or slot blot may also be used to arrange and.link
elements to the surface
of a substrate using thermal, UV, chemical, or mechanical bonding procedures.
A typical array may
be produced using available methods and machines well known to those of
ordinary skill in the art
and may contain any appropriate number of elements (Schena, M. et al. (1995)
Science 270:467-470;
Shalom D. et al. (1996) Genome Res. 6:639-645; Marshall, A. and J. Hodgson
(1998) Nat.
Biotechnol. 16:27-31).
Full length cDNAs, Expressed Sequence Tags (ESTs), or fragments or oligomers
thereof may
comprise the elements of the microarray. Fragments or oligomers suitable for
hybridization can be
selected using software well known in the art such as LASERGENE software
(DNASTAR). The
array elements are hybridized with polynucleotides in a biological sample. The
polynucleotides in
the biological sample are conjugated to a fluorescent label or other molecular
tag for ease of
detection. After hybridization, nonhybridized nucleotides from the biological
sample are removed,
and a fluorescence scanner is used to detect hybridization at each array
element. Alternatively, laser
desorbtion and mass spectrometry may be used for detection of hybridization.
The degree of
complementarity and the relative abundance of each polynucleotide which
hybridizes to an element
on the microarray may be assessed. In one embodiment, microarray preparation
and usage is
described in detail below.
Tissue or Cell Sample Preparation
Total RNA is isolated from tissue samples using the guanidinium thiocyanate
method and
poly(A)~ RNA is purified using the oligo-(dT) cellulose method. Each poly(A)+
RNA sample is
reverse transcribed using MMLV reverse-transcriptase, 0.05 pg/~1 oligo-(dT)
primer (2lmer), 1X
first strand buffer, 0.03 unitsl~.l RNase inhibitor, 500 ~,M dATP, 500 p.M
dGTP, 500 ~M dTTP, 40
~,M dCTP, 40 ~.M dCTP-Cy3 (BDS) or dCTP-Cy5 (Amersham Biosciences). The
reverse
transcription reaction is performed in a 25 ml volume containing 200 ng
poly(A)+ RNA with
GEMBRIGHT kits (Incyte Genomics). Specific control poly(A)~ RNAs are
synthesized by izz vitro
transcription from non-coding yeast genomic DNA. After incubation at 37
° C for 2 hr, each reaction
sample (one with Cy3 and another with Cy5 labeling) is treated with 2.5 ml of
0.5M sodium
hydroxide and incubated for 20 minutes at 85° C to the stop the
reaction and degrade the RNA.
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Samples are purified using two successive CHROMA SPIN 30 gel filtration spin
columns (Clontech,
Palo Alto CA) and after combining, both reaction samples are ethanol
precipitated using 1 ml of
glycogen (1 mg/ml), 60 ml sodium acetate, and 300 ml of 100% ethanol. The
sample is then dried to
completion using a SpeedVAC (Savant Instruments Inc., Holbrook NY) and
resuspended in 14 ~,15X
SSC/0.2% SDS.
Microarray Preparation
Sequences of the present invention are used to generate array elements. Each
array element
is amplified from bacterial cells containing vectors with cloned cDNA inserts.
PCR amplification
uses primers complementary to the vector sequences flanking the cDNA insert.
Array elements are
amplified in thirty cycles of PCR from an initial quantity of 1-2 ng to a
final quantity greater than 5
~,g. Amplified array elements are then purified using SEPHACRYL-400 (Amersham
Biosciences).
Purified array elements are immobilized on polymer-coated glass slides. Glass
microscope
slides (Corning) are cleaned by ultrasound in 0.1% SDS and acetone, with
extensive distilled water
washes between and after treatments. Glass slides are etched in 4%
hydrofluoric acid (VWR
Scientific Products Corporation (VWR), West Chester PA), washed extensively in
distilled water,
and coated with 0.05% aminopropyl silane (Sigma) in 95% ethanol. Coated slides
are cured in a
110°C oven.
Array elements are applied to the coated glass substrate using a procedure
described in U.S.
Patent No. 5,807,522, incorporated herein by reference. 1 ~,l of the array
element DNA, at an average
concentration of 100 ng/~.1, is loaded into the open capillary printing
element by a high-speed robotic
apparatus. The apparatus then deposits about 5 n1 of array element sample per
slide.
Microarrays are W-crosslinked using a STRATALINKER UV-crosslinker
(Stratagene).
Microarrays are washed at room temperature once in 0.2% SDS and three times in
distilled water.
Non-specific binding sites are blocked by incubation of microarrays in 0.2%
casein in phosphate
buffered saline (PBS) (Tropix, Inc., Bedford MA) for 30 minutes at 60°C
followed by washes in
0.2% SDS and distilled water as before.
Hybridization
Hybridization reactions contain 9 ~,l of sample mixture consisting of 0.2 ~,g
each of Cy3 and
Cy5 labeled cDNA synthesis products in 5X SSC, 0.2% SDS hybridization buffer.
The sample
mixture is heated to 65° C for 5 minutes and is aliquoted onto the
microarray surface and covered
with an 1.8 cm' coverslip. The arrays are transferred to a waterproof chamber
having a cavity just
slightly larger than a microscope slide. The chamber is kept at 100% humidity
internally by the
addition of 140 ~.1 of 5X SSC in a corner of the chamber. The chamber
containing the arrays is
incubated for about 6.5 hours at 60° C. The arrays are washed for 10
min at 45° C in a first wash
buffer (1X SSC, 0.1% SDS), three times for 10 minutes each at 45°C in a
second wash buffer (0.1X
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SSC), and dried.
Detection
Reporter-labeled hybridization complexes are detected with a microscope
equipped with an
Innova 70 mixed gas 10 W laser (Coherent, Inc., Santa Clara CA) capable of
generating spectral lines
at 488 nm for excitation of Cy3 and at 632 nm for excitation of CyS. The
excitation laser light is
focused on the array using a 20X microscope objective (Nikon, Inc., Melville
NY). The slide
containing the array is placed on a computer-controlled X-Y stage on the
microscope and raster-
scanned past the objective. The 1.8 cm x 1.8 cm array used in the present
example is scanned with a
resolution of 20 micrometers.
In two separate scans, a mixed gas multiline laser excites the two
fluorophores sequentially.
Emitted light is split, based on wavelength, into two photomultiplier tube
detectors (PMT 81477,
Hamamatsu Photonics Systems, Bridgewater NJ) corresponding to the two
fluorophores.
Appropriate filters positioned between the array and the photomultiplier tubes
are used to filter the
signals. The emission maxima of the fluorophores used are 565 nm for Cy3 and
650 nm for CyS.
Each array is typically scanned twice, one scan per fluorophore using the
appropriate filters at the
laser source, although the apparatus is capable of recording the spectra from
both fluorophores
simultaneously.
The sensitivity of the scans is typically calibrated using the signal
intensity generated by a
cDNA control species added to the sample mixture at a known concentration. A
specific location on
the array contains a complementary DNA sequence, allowing the intensity of the
signal at that
location to be correlated with a weight ratio of hybridizing species of
1:100,000. When two samples
from different sources (e.g., representing test and control cells), each
labeled with a different
fluorophore, are hybridized to a single array for the purpose of identifying
genes that are
differentially expressed, the calibration is done by labeling samples of the
calibrating cDNA with the
two fluorophores and adding identical amounts of each to the hybridization
mixture.
The output of the photomultiplier tube is digitized using a 12-bit RTI-835H
analog-to-digital
(AiD) conversion board (Analog Devices, Inc., Norwood MA) installed in an IBM-
compatible PC
computer. The digitized data are displayed as an image where the signal
intensity is mapped using a
linear 20-color transformation to a pseudocolor scale ranging from blue (low
signal) to red (high
signal). The data is also analyzed quantitatively. Where two different
fluorophores are excited and
measured simultaneously, the data are first corrected for optical crosstalk
(due to overlapping
emission spectra) between the fluorophores using each fluorophore's emission
spectrum.
A grid is superimposed over the fluorescence signal image such that the signal
from each
spot is centered in each element of the grid. The fluorescence signal within
each element is then
integrated to obtain a numerical value corresponding to the average intensity
of the signal. The
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software used for signal analysis is the GEMTOOLS gene expression analysis
program (Incyte
Genomics). Array elements that exhibit at least about a two-fold change in
expression, a signal-to-
background ratio of at least about 2.5, and an element spot size of at least
about 40%, are considered
to be differentially expressed.
Expression
SEQ ID N0:58 showed differential expression in mild Alzheimer's Disease as
determined by
microarray analysis. Alzheimer's Disease (AD) is a progressive dementia
characterized
neuropathologically by the presence of amyloid beta-peptide-containing plaques
and neurofibrillary
tangles in specific brain regions. In addition, neurons and synapses are lost
and inflammatory
responses are activated in microglia and astrocytes. A cross comparison of
normal posterior cingulate
brain tissue to posterior cingulate brain tissue showing mild AD was carried
out. SEQ ID NO:58
showed at least two-fold increased expression in the tissue of a 68-year-old
female donor with mild
AD, as compared to the tissue from a 61-year-old female donor with no AD. This
experiment
indicates that SEQ D7 N0:58 is useful in diagnostic assays and disease staging
for AD and as a
potential biological marker and therapeutic agent in the treatment of AD.
For example, SEQ ID N0:82 showed differential expression in breast cancer
tissue, as
determined by microarray analysis. Histological and molecular evaluation of
breast tumors has
revealed that the development of breast cancer evolves through a mufti-step
process whereby pre-
malignant mammary epithelial cells undergo a relatively defined sequence of
events leading to tumor
formation. Early in tumor development ductal hyperplasia is observed. Cells
undergoing rapid
neoplastic growth gradually progress to invasive carcinoma and become
metastatic to the lung, bone
and potentially other organs. Several factors, ranging from, but not limited
to, environmental to
genetic, influence tumor progression and malignant transformation. In order to
better determine the
molecular and phenotypic characteristics associated with different stages of
breast cancer, breast
carcinoma cell lines at various stages of tumor progression were compared to
primary human breast
epithelial cells. MDA-mb-231, a breast tumor cell line isolated from the
pleural effusion of a 51-
year-old female, which forms poorly differentiated adenocarcinoma in nude mice
and expresses the
Wnt3 oncogene, EGF and TGF-a, was compared to two non-cancerous cell lines,
HMEC and MCF-
10A. The primary mammary epithelial cell line HMEC was derived from normal
human mammary
tissue (Clonetics, San Diego, CA). MCF-l0A is a breast mammary gland cell line
isolated from a 36-
year-old female with fibrocystic disease. The expression of SEQ 117 N0:82 was
increased by at least
two-fold in MDA-mb-231 cells relative to HMEC and MCF-l0A cells. Therefore,
SEQ ID N0:82
can be useful in diagnostic and staging assays for breast cancer and as a
potential biological marker
and therapeutic agent in the treatment of breast cancer.
In another example, SEQ ID NO:86 showed differential expression, as determined
by
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microarray analysis, in inflammatory responses. Human peripheral blood
mononuclear cells
(PBMCs) (52% lymphocytes, 20% NK cells, 25% monocytes, and 3% various cells
that include
dendritic and progenitor cells) were treated with the pro-inflammatory
cytokines interleukin-1 (3,
interleukin-2, interleukin-6, interleukin-8, interleukin-12, interleukin-18,
tumor necrosis factor-cc and
interferon-'y, for 2 and 4 hours. The expression of SEQ )D N0:86 was increased
by at least two-fold
at both time points, as compared to untreated PBMCs. Therefore, SEQ ID N0:86
is useful in
diagnostic assays for inflammatory responses and as a potential biological
marker and therapeutic
agent in the treatment of inflammatory responses.
SEQ >D N0:86 also showed differential expression in Alzheimer's Disease (AD),
as
determined by microarray analysis. AD is a progressive neurodegenerative
disorder that is
characterized by the formation of senile plaques and neurofibrillary tangles
containing amyloid beta
peptide. These plaques are found in limbic and association cortices of the
brain. The hippocampus is
part of the limbic system and plays an important role in learning and memory.
In subjects with AD,
accumulating plaques damage the neuronal architecture in limbic areas and
eventually cripple the
memory process. In a comparison of cingulate posterior brain tissue from a 68-
year-old female with
mild AD to anterior hippocampal tissue from a normal 61-year-old female, the
expression of SEQ ID
N0:86 was increased at least four-fold. Therefore, SEQ )D N0:86 is useful in
diagnostic assays for
AD and as a potential biological marker and therapeutic agent in the treatment
of AD.
For example. SEQ ID NO:96 showed differential expression in certain prostate
carcinoma
cell lines versus normal prostate epithelial cells as determined by microarray
analysis. The prostate
carcinoma cell lines include DU 145, LNCaP, and PC-3. DU 145 was isolated from
a metastatic site
in the brain of a 69 year old male with widespread metastatic prostate
carcinoma. DU 145 has no
detectable sensitivity to hormones; forms colonies in semi-solid medium; is
only weakly positive for
acid phosphatase; and cells are negative for prostate specific antigen (PSA).
LNCaP is a prostate
carcinoma cell line isolated from a lymph node biopsy of a 50 year old male
with metastatic prostate
carcinoma. LNCaP expresses PSA, produces prostate acid phosphatase, and
expresses androgen
receptors. PC-3, a prostate adenocarcinoma cell line, was isolated from a
metastatic site in the bone
of a 62 year old male with grade IV prostate adenocarcinoma. The normal
epithelial cell line, PrEC,
is a primary prostate epithelial cell line isolated from a normal donor. This
experiment showed that
the expression of SEQ ID N0:96 was decreased by at least two fold in both DU
145 and LNCaP cells
compared to PrEC cells. Therefore, SEQ )I? N0:96 is useful as a diagnostic
marker or as a potential
therapeutic target for certain prostate cancers.
XII. Complementary Polynucleotides
Sequences complementary to the MDDT-encoding sequences, or any parts thereof,
are used
to detect, decrease, or inhibit expression of naturally occurring MDDT.
Although use of
~s
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oligonucleotides comprising from about 15 to 30 base pairs is described,
essentially the same
procedure is used with smaller or with larger sequence fragments. Appropriate
oligonucleotides are
designed using OLIGO 4.06 software (National Biosciences) and the coding
sequence of MDDT. To
inhibit transcription, a complementary oligonucleotide is designed from the
most unique 5' sequence
and used to prevent promoter binding to the coding sequence. To inhibit
translation, a
complementary oligonucleotide is designed to prevent ribosomal binding to the
MDDT-encoding
transcript.
XIII. Expression of MDDT
Expression and purification of MDDT is achieved using bacterial or virus-based
expression
systems. For expression of MDDT in bacteria, cDNA is subcloned into an
appropriate vector
containing an antibiotic resistance gene and an inducible promoter that
directs high levels of cDNA
transcription. Examples of such promoters include, but are not limited to, the
trp-lac (tac) hybrid
promoter and the T5 or T7 bacteriophage promoter in conjunction with the lac
operator regulatory
element. Recombinant vectors are transformed into suitable bacterial hosts,
e.g., BL21(DE3).
Antibiotic resistant bacteria express MDDT upon induction with isopropyl beta-
D-
thiogalactopyranoside (IPTG). Expression of MDDT in eukaryotic cells is
achieved by infecting
insect or mammalian cell lines with recombinant Autographica californiea
nuclear polyhedrosis virus
(AcMNPV), commonly known as baculovirus. The nonessential polyhedrin gene of
baculovirus is
replaced with cDNA encoding MDDT by either homologous recombination or
bacterial-mediated
transposition involving transfer plasmid intermediates. Viral infectivity is
maintained and the strong
polyhedrin promoter drives high levels of cDNA transcription. Recombinant
baculovirus is used to
infect Spodopter-a frugiperda (Sf9) insect cells in most cases, or human
hepatocytes, in some cases.
Infection of the latter requires additional genetic modifications to
baculovirus (Engelhard, E.K. et al.
(1994) Proc. Natl. Acad. Sci. USA 91:3224-3227; Sandig, V. et al. (1996) Hum.
Gene Ther. 7:1937-
1945).
In most expression systems, MDDT is synthesized as a fusion protein with,
e.g., glutathione
S-transferase (GST) or a peptide epitope tag, such as FLAG or 6-His,
permitting rapid, single-step,
affinity-based purification of recombinant fusion protein from crude cell
lysates. GST, a 26-
kilodalton enzyme from Schistosoma japonicur~i, enables the purification of
fusion proteins on
immobilized glutathione under conditions that maintain protein activity and
antigenicity (Amersham
Biosciences). Following purification, the GST moiety can be proteolytically
cleaved from MDDT at
specifically engineered sites. FLAG, an 8-amino acid peptide, enables
immunoaffinity purification
using commercially available monoclonal and polyclonal anti-FLAG antibodies
(Eastman Kodak). 6-
His, a stretch of six consecutive histidine residues, enables purification on
metal-chelate resins
(QIAGEN). Methods for protein expression and purification are discussed in
Ausubel et al. (supra,
79
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
ch. 10 and 16). Purified MDDT obtained by these methods can be used directly
in the assays shown
in Examples XVII and XVIII, where applicable.
XIV. Functional Assays
MDDT function is assessed by expressing the sequences encoding MDDT at
physiologically
elevated levels in mammalian cell culture systems. cDNA is subcloned into a
mammalian expression
vector containing a strong promoter that drives high levels of cDNA
expression. Vectors of choice
include PCMV SPORT plasmid (Invitrogen, Carlsbad CA) and PCR3.1 plasmid
(Invitrogen), both of
which contain the cytomegalovirus promoter. 5-10 ~g of recombinant vector are
transiently
transfected into a human cell line, for example, an endothelial or
hematopoietic cell line, using either
liposome formulations or electroporation. 1-2 ,ug of an additional plasmid
containing sequences
encoding a marker protein are co-transfected. Expression of a marker protein
provides a means to
distinguish transfected cells from nontransfected cells and is a reliable
predictor of cDNA expression
from the recombinant vector. Marker proteins of choice include, e.g., Green
Fluorescent Protein
(GFP; Clontech), CD64, or a CD64-GFP fusion protein. Flow cytometry (FCM), an
automated, laser
optics-based technique, is used to identify transfected cells expressing GFP
or CD64-GFP and to
evaluate the apoptotic state of the cells and other cellular properties. FCM
detects and quantifies the
uptake of fluorescent molecules that diagnose events preceding or coincident
with cell death. These
events include changes in nuclear DNA content as measured by staining of DNA
with propidium
iodide; changes in cell size and granularity as measured by forward light
scatter and 90 degree side
light scatter; down-regulation of DNA synthesis as measured by decrease in
bromodeoxyuridine
uptake; alterations in expression of cell surface and intracellular proteins
as measured by reactivity
with specific antibodies; and alterations in plasma membrane composition as
measured by the binding
of fluorescein-conjugated Annexin V protein to the cell surface. Methods in
flow cytometry are
discussed in Ormerod, M.G. (1994; Flow Cytometry, Oxford, New York NY).
The influence of MDDT on gene expression can be assessed using highly purified
populations of cells transfected with sequences encoding MDDT and either CD64
or CD64-GFP.
CD64 and CD64-GFP are expressed on the surface of transfected cells and bind
to conserved regions
of human immunoglobulin G (IgG). Transfected cells are efficiently separated
from nontransfected
cells using magnetic beads coated with either human IgG or antibody against
CD64 (DYNAL, Lake
Success NY). mRNA can be purified from the cells using methods well known by
those of skill in
the art. Expression of mRNA encoding MDDT and other genes of interest can be
analyzed by
northern analysis or microarray techniques.
XV. Production of MDDT Specific Antibodies
MDDT substantially purified using polyacrylamide gel electrophoresis (PAGE;
see, e.g.,
Harrington, M.G. ( 1990) Methods Enzymol. 182:488-495), or other purification
techniques, is used to
so
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
immunize animals (e.g., rabbits, mice, etc.) and to produce antibodies using
standard protocols.
Alternatively, the MDDT amino acid sequence is analyzed using LASERGENE
software
(DNASTAR) to determine regions of high immunogenicity, and a corresponding
oligopeptide is
synthesized and used to raise antibodies by means known to those of skill in
the art. Methods for
selection of appropriate epitopes, such as those near the C-terminus or in
hydrophilic regions are well
described in the art (Ausubel et al., supra, ch. 11).
Typically, oligopeptides of about 15 residues in length are synthesized using
an ABI 431A
peptide synthesizer (Applied Biosystems) using FMOC chemistry and coupled to
KLH (Sigma-
Aldrich, St. Louis MO) by reaction with N-maleimidobenzoyl-N-
hydroxysuccinimide ester (MBS) to
increase immunogenicity (Ausubel et al., supra). Rabbits are immunized with
the oligopeptide-KLH
complex in complete Freund's adjuvant. Resulting antisera are tested for
antipeptide and anti-MDDT
activity by, for example, binding the peptide or MDDT to a substrate, blocking
with 1% BSA,
reacting with rabbit antisera, washing, and reacting with radio-iodinated goat
anti-rabbit IgG.
XVI. Purification of Naturally Occurring MDDT Using Specific Antibodies
Naturally occurring or recombinant MDDT is substantially purified by
immunoaffmity
chromatography using antibodies specific for MDDT. An immunoaffinity column is
constructed by
covalently coupling anti-MDDT antibody to an activated chromatographic resin,
such as
CNBr-activated SEPHAROSE (Amersham Biosciences). After the coupling, the resin
is blocked and
washed according to the manufacturer's instructions.
Media containing MDDT are passed over the immunoaffinity column, and the
column is
washed under conditions that allow the preferential absorbance of MDDT (e.g.,
high ionic strength
buffers in the presence of detergent). The column is eluted under conditions
that disrupt
antibody/MDDT binding (e.g., a buffer of pH 2 to pH 3, or a high concentration
of a chaotrope, such
as urea or. thiocyanate ion), and MDDT is collected.
XVII. Identification of Molecules Which Interact with MDDT
MDDT, or biologically active fragments thereof, are labeled with'zsI Bolton-
Hunter reagent
(Bolton, A.E. and W.M. Hunter (1973) Biochem. J. 133:529-539). Candidate
molecules previously
arrayed in the wells of a mufti-well plate are incubated with the labeled
MDDT, washed, and any
wells with labeled MDDT complex are assayed. Data obtained using different
concentrations of
MDDT are used to calculate values for the number, affinity, and association of
MDDT with the
candidate molecules.
Alternatively, molecules interacting with MDDT are analyzed using the yeast
two-hybrid
system as described in Fields, S. and O. Song (1989; Nature 340:245-246), or
using commercially
available kits based on the two-hybrid system, such as the MATCHMAKER system
(Clontech).
MDDT may also be used in the PATHCALLING process (CuraGen Corp., New Haven CT)
81
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
which employs the yeast two-hybrid system in a high-throughput manner to
determine all interactions
between the proteins encoded by two large libraries of genes (Nandabalan, I~.
et al. (2000) U.S.
Patent No. 6,057,101).
XVIII. Demonstration of MDDT Activity
Phorbol ester binding activity of MDDT is measured using an assay based on the
fluorescent
phorbol ester sapinotoxin-D (SAPD). Binding of SAPD to MDDT is quantified by
measuring the
resonance energy transfer from MDDT tryptophans to the 2-(N-
methylamino)benzoyl fluorophore of
the phorbol ester, as described by Slater et al. ((1996) J. Biol. Chem.
271:4627-4631).
Various modifications and variations of the described compositions, methods,
and systems of
the invention will be apparent to those skilled in the art without departing
from the scope and spirit of
the invention. It will be appreciated that the invention provides novel and
useful proteins, and their
encoding polynucleotides, which can be used in the drug discovery process, as
well as methods for
using these compositions for the detection, diagnosis, and treatment of
diseases and conditions.
Although the invention has been described in connection with certain
embodiments, it should be
understood that the invention as claimed should not be unduly limited to such
specific embodiments.
Nor 'should the description of such embodiments be considered exhaustive or
limit the invention to
the precise forms disclosed. Furthermore, elements from one embodiment can be
readily recombined
with elements from one or more other embodiments. Such combinations can form a
number of
embodiments within the scope of the invention. It is intended that the scope
of the invention be
defined by the following claims and their equivalents.
82
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
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Table 5
Polynucleotidencyte ProjectRepresentative Library
SEQ I ID:
ID NO:
49 1629602CB COLNPOT01
1
50 2100360CB1 BRAHNON05 .
51 5166833CB LUNGNOT31
1
52 7494963CB NOSETUE01
1
53 7644881CB1 NERDTDN03
54 3790383CB1 BRSTNOT28
55 3846110CB DENDNOTO1
1
56 1878279CB1 ENDMUNE01
57 1848891CB OVARNOT03
1
58 2500251CB1 BRAVUNT02
59 55026561CB1LUNGDIS03
60 7502593CB BRANDINO1
1
61 7503957CB OVARTUT04
1
62 7504415CB1 THP1NOB01
63 7504074CB NEUTFMTOl
1
64 7502257CB MCLRNOCO1
1
65 1315136CB LUNGNOT09
1
66 1379785CB1 LUNGNOT10
67 2011166CB TESTNOT03
1
68 3434684CB OVARDIRO1
1
69 5134056CB1 PROSNOT14
70 5281724CB ADRETUROl
1
71 7502391CB1 LUNGNOT38
72 7502544CB KIDNTUE01
1
73 2858465CB1 DRGCNOTOl
74 7503455CB BRAENOT04
1
75 7503479CB1 293TF2T01
76 7218127CB SINTNOR01
1
77 1688943CB1 THP1NOT03
78 2369350CB BRAITUT02
1
79 2722979CB HNT2AZS07
1
80 60140470CB MI~7UNB01
1
81 70623603CB BRSTUNFO1
1
82 7161479CB LIVRNON08
1
83 7502313CB1 BONRFETOl
84 7502390CB THYRNOT03
1
85 7502872CB1 LUNGTUT08
86 7505443CB1 293TF2T01
87 8032443CB TESTNOFO1
1
88 7704916CB TESTNOT03
1
89 2013440CB LUNGASTOl
1
90 2503512CB CONUTUTO1
1
91 277396CB TESTNOT03
1
92 3044046CB TONSDIE01
1 ~
93 3808420CB1 LUNGNOT04
94 7504028CB BRSTTUT08
1
95 7766880CB BRAINONO1
1
96 90089609CB LUNGTUT08
1
11G
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CA 02460480 2004-03-15
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CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
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127
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<110> INCYTE GENOMICS. INC.
TANG, Y. Tom
FORSYTHE, Ian J.
EMERLING, Brooke M.
HAFALIA, April J.A.
YUE, Henry
XU, Yuming
GIETZEN, Kimberly J.
CHAWLA, Narinder K.
BAUGHN, Mariah R.
MARQUIS, Joseph P.
BECHA, Shanya D.
KABLE, Amy E.
LAL, Preeti G.
RICHARDSON, Thomas W.
LEE, Soo Yeun
LEE, Ernestine A.
TRAM, Bao
WARREN, Bridget A.
LU, Dyung Aina M.
GURURAJAN, Rajagopal
SPRAGUE, William W.
BLAKE, Julie J.
THANGAVELU, Kavitha
SWARNAKAR, Anita
GORVAD, Ann E.
GRTFFIN, Jennifer A.
LINDQUIST, Erika A.
ELLIOTT, Vicki S.
ISON, Craig H.
RAMKUMAR, Jayala~ni
<120> MOLECULES FOR DISEASE DETECTION AND TREATMENT
<130> PF-1232 PCT
<140> To Be Assigned
<141> Herewith
<150> US 60/328,944
<151> 2001-10-12
<150> US 60/345,384
<151> 2001-10-26
<150> US 60/343,880
<151> 2001-11-02
<150> US 60/345,143
<151> 2001-11-09
<150> US 60/332,430
<151> 2001-11-16
<160> 96
<170> PERL Program
<210> 1
<211> 93
<212> PRT
<213> Homo sapiens
<220>
1/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<221> misc_feature
<223> Incyte ID No: 1629602CD1
<400> 1
Met Val Gln Ala Gly Pro Ser Ser Cys Ser Ile Ser Gly Asp Pro
1 5 10 15
Gly Leu Pro Arg Arg Trp Arg Pro Ala Gln Va1 Val Arg Pro Gly
20 25 30
Arg Leu Arg Ile Arg Gly Trp Ser Arg Arg Ile Pro Lys Ala Glu
35 40 45
Val Gly Ser Pro Gly Asp Ser Gln Leu Leu Ser Leu Trp Arg Arg
50 55 60
Gly Pro Val Thr Glu Ala Pro Phe Ser Asn Pro Gly Ala Ala Phe
65 70 75
Ser Pro Tyr Arg Lys Ser Asp Gly Leu Met Thr Ser Trp Leu Ala
80 85 90
Ala Glu Arg
<210> 2
<211> 281
<212> PRT
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 2100360CD1
<400> 2
Met I1e Leu Thr Lys A1a Gln Tyr Asp Glu Ile Ala Gln Cys Leu
1 5 10 15
Val Ser Val Pro Pro Thr Arg Gln Ser Leu Arg Lys Leu Lys Gln
20 25 30
Arg Phe Pro Ser Gln Ser Gln Ala Thr Leu Leu Ser Ile Phe Ser
35 40 45
Gln Glu Tyr G1n Lys His Ile Lys Arg Thr His Ala Lys His His
50 55 60
Thr Ser Glu A1a Ile G1u Ser Tyr Tyr Gln Arg Tyr Leu Asn Gly
65 70 75
Val Val Lys Asn Gly Ala Ala Pro Val Leu Leu Asp Leu A1a Asn
8D 85 90
G1u Val Asp Tyr Ala Pro Ser Leu Met Ala Arg Leu Ile Leu Glu
95 100 105
Arg Phe Leu G1n Glu His Glu G1u Thr Pro Pro Ser Lys Ser Ile
110 115 120
Ile Asn Ser Met Leu Arg Asp Pro Ser G1n Ile Pro Asp Gly Val
125 130 135
Leu Ala Asn Gln Val Tyr Gln Cys Ile Val Asn Asp Cys Cys Tyr
140 145 150
Gly Pro Leu Val Asp Cys Ile Lys His A1a Ile Gly His Glu His
155 160 165
Glu Val Leu Leu Arg Asp Leu Leu Leu Glu Lys Asn Leu Ser Phe
170 175 180
Leu Asp Glu Asp Gln Leu Arg Ala Lys Gly Tyr Asp Lys Thr Pro
185 190 195
Asp Phe Ile Leu Gln Val Pro Val Ala Val G1u Gly His Ile Ile
200 205 210
His Trp Ile Glu Ser Lys Ala Ser Phe Gly Asp G1u Cys Ser His
215 220 225
His Ala Tyr Leu His Asp Gln Phe Trp Ser Tyr Trp Asn Arg Phe
230 ~ 235 240
Gly Pro Gly Leu Val Ile Tyr Trp Tyr Gly Phe Ile Gln Glu Leu
245 250 255
2/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
Asp Cys Asn Arg Glu Arg Gly Ile Leu Leu Lys Ala Cys Phe Pro
260 265 270
Thr Asn Ile Val Thr Leu Cys His Ser Ile Ala
275 280
<210> 3
<211> 292
<212> PRT
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 5166833CD1
<400> 3
Met Ser Ile Ser Leu Ser Ser Leu Ile Leu Leu Pro Ile Trp Ile
1 5 10 15
Asn Met Ala Gln Ile Gln Gln Gly Gly Pro Asp Glu Lys Glu Lys
20 25 30
Thr Thr Ala Leu Lys Asp Leu Leu Ser Arg Ile Asp Leu Asp Glu
35 40 45
Leu Met Lys Lys Asp Glu Pro Pro Leu Asp Phe Pro Asp Thr Leu
50 55 60
Glu Gly Phe Glu Tyr Ala Phe Asn Glu Lys Gly Gln Leu Arg His
65 70 75
Ile Lys Thr Gly Glu Pro Phe Val Phe Asn Tyr Arg Glu Asp Leu
80 85 90
His Arg Trp Asn Gln Lys Arg Tyr Glu Ala~Leu Gly Glu Ile Ile
95 100 105
Thr Lys Tyr Val Tyr Glu Leu Leu Glu Lys Asp Cys Asn Leu Lys
110 115 120
Lys Val Ser Ile Pro Val Asp A1a Thr G1u Ser Glu Pro Lys Ser
125 130 135
Phe Ile Phe Met Ser Glu Asp A1a Leu Thr Asn Pro Gln Lys Leu
140 145 150
Met Va1 Leu Ile His Gly Ser Gly Va1 Val Arg Ala Gly Gln Trp
155 160 165
Ala Arg Arg Leu Ile Ile Asn Glu Asp Leu Asp Ser Gly Thr Gln
170 175 180
Ile Pro Phe Ile Lys Arg Ala Val Ala G1u Gly Tyr Gly Val Ile
185 190 195
Val Leu Asn Pro Asn Glu Asn Tyr Ile Glu Val Glu Lys Pro Lys
200 205 210
Ile His Val Gln Ser Ser Ser Asp Ser Ser Asp Glu Pro Ala Glu
215 220 225
Lys Arg Glu Arg Lys Asp Lys Val Ser Lys Glu Thr Lys Lys Arg
230 235 240
Arg Asp Phe Tyr Glu Lys Tyr Arg Asn Pro Gln Lys Lys Lys Glu
245 250 255
Met Met Gln Leu Tyr Ile Arg Val Ser G1u Ile Thr Thr Phe Leu
260 265 270
Tyr Tyr Phe Leu Tyr Leu Val Tyr Ile Leu Leu Tyr Val Asp Cys
275 280 285
Phe Val Phe Leu Gln Glu Tyr °
290
<210> 4
<211> 270
<212> PRT
<213> Homo sapiens
<220>
<221> misc_feature
3190
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<223> Incyte ID No: 7494963CD1
<400> 4
Met Glu Ala Glu Glu Leu Glu Gln Glu Arg Glu Gln Leu Arg Leu
1 5 10 15
Gln Leu Trp G1u A1a Tyr Cys Gln Val Arg Tyr Leu Cys Ser His
20 25 30
Leu Arg Gly Asn Asp Ser Ala Asp Ser Ala Val Ser Thr Asp Ser
35 40 45
Ser Met Asp Glu Ser Ser Glu Thr Ser Ser Ala Lys Asp Val Pro
50 55 60
Ala Gly Ser Leu Arg Thr Ala Leu Asn Glu Leu Lys Arg Leu Ile
65 70 75
Gln Ser Ile Val Asp G1y Met Glu Pro Thr Gly Ser Arg Arg Leu
80 85 90
Asp Asp Asp Ser Leu Glu Glu Gln Ile Arg Gln Thr Ser Glu Asp
95 100 105
Ser Arg Ala Leu Arg Glu Leu Met Glu Gly Glu Arg Gly Lys Leu
110 115 120
Arg Gln Ser Leu Glu Glu Leu Gln Arg Leu His Ser Gln Val Thr
125 130 135
Leu Leu Ser Val Glu Met Thr Ala Leu Lys Glu Glu Arg Asp Arg
140 145 150
Leu Arg Val Thr Ser Glu Asp Lys Glu Pro Lys Glu Gln Leu Gln
155 160 165
Lys Ala Ile Arg Asp Arg Asp Glu A1a Ile Ala Lys Lys Asn Ala
170 175 180
Val Glu Leu G1u Leu Ala Lys Cys Arg Met Asp Met Met Ser Leu
185 190 195
Asn Ser Gln Leu Leu Asp Ala I1e Gln Gln Lys Leu Asn Leu Ser
200 205 210
Gln Gln Leu Glu Ala Trp Gln Asp Asp Met His Arg Val I1e Asp
215 220 225
Arg G1n Leu Met Asp Thr His Leu Lys Glu Arg Ser Gln Pro Ala
230 235 240
Ala Ala Leu Cys Arg Gly His Ser Ala Gly Arg Gly Asp Glu Pro
245 250 255
Ser I1e Ala Glu Gly Lys Arg Leu Phe Ser Phe Phe Arg Lys Ile
260 265 270
<210> 5
<211> 447
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7644881CD1
<400> 5
Met Gln Glu Ser G1n Glu Thr His Ile Ser Asn His Leu Asp Glu
1 5 10 15
Val Val Ala Ala Val Ser Ile Thr His Arg Lys Lys Phe Gln Asn
20 ~ 25 30
Lys Leu Leu Gln Thr Ala Leu Phe Gln Pro Pro Arg Glu Lys Leu
35 40 45
His Leu Cys Glu Glu Lys Ala Lys Ser Tyr Ser Asn Ser His Glu
50 55 60
Tyr Lys G1n A1a Val His Glu Leu Val Arg Cys Val Ala Leu Thr
65 70 75
Arg Ile Cys Tyr Gly Asp Ser His Trp Lys Leu Ala Glu A1a His
80 85 90
4/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
Va1 Asn Leu Ala Gln Gly Tyr Leu Gln Leu Lys G1y Leu Ser Leu
95 100 105
G1n Ala Lys G1n His Ala Glu Lys Ala Arg Gln Ile Leu A1a Asn
110 115 120
Ser Ile Val Pro Pro Tyr Ser Glu Asn Thr Asp Val Phe Lys Phe
125 130 135
Ser Ile G1u Leu Phe His Thr Met Gly Arg Ala Leu Leu Ser Leu
140 145 150
Gln Lys Phe Lys Glu Ala Ala Glu Asn Leu Thr Lys Ala Glu Arg
155 160 165
Leu Ser Lys Glu Leu Leu' Gln Cys Gly Arg I1e Ile Lys Glu Glu
170 175 180
Trp Ile Glu Ile Glu A1a Arg Ile Arg Leu Ser Phe Ala Gln Val
185 190 195
Tyr Gln Gly Gln Lys Lys Ser Lys Glu Ala Leu Ser His Tyr Gln
200 205 210
A1a Ala Leu Glu Tyr Val Glu Ile Ser Lys Gly Glu Thr Ser Arg
215 220 225
Glu Cys Val Pro Ile Leu Arg Glu Leu Ala G1y Val Glu Gln Ala
230 235 240
Leu Gly Leu His Asp Val Ser Ile Asn His Phe Leu Gln Ala His
245 250 255
Leu Tle Ile Leu Ser Arg Ser Pro Ser Gln Val Glu Ala Ala Asp
260 265 270
Ser Ala His Ile Val Ala His Ala Ala Val Ala Ser Gly Arg His
275 280 285
Glu His His Asp Val Ala Glu Gln Tyr Phe Gln Glu Ser Met Ala
290 295 300
His Leu Lys Asp Ser Glu Gly Met Gly Arg Thr Lys Phe Leu Ser
305 310 315
Ile Gln Asp Glu Phe Cys His Phe Leu Gln Met Thr Gly Gln Lys
320 325 330
Glu Arg Ala Thr Ser Ile Leu Arg Glu Ser Leu Glu Ala Lys Val
335 340 345
Glu A1a Phe Gly Asp Phe Ser Pro Glu Val Ala Glu Thr Tyr Arg
350 355 360
Leu Leu Gly Gly Ala Asp Leu Ala Gln Gly Asn His Ser Gly A1a
365 370 375
Arg Lys Lys Leu Lys Lys Cys Leu Gln Ile Gln Thr Leu Leu Tyr
380 385 390
Gly Pro Gln Asp Lys Arg Thr Leu Ala Thr Gln Gln Ala Met Gly
395 400 405
Met Leu Ser Thr Ala Pro Lys Val Ala Ser Lys Pro Arg Gln Ala
410 415 420
Ser Lys Ala Lys Val Ala Phe Cys Thr Ser Ile Pro Gln Asp Thr
425 430 435
Leu Leu Gly Lys Ala Arg Pro Gly Thr Thr Ala Asp
440 445
<210> 6
<211> 757
<212> PRT
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 3790383CD1
<400> 6
Met A1a Glu Val Gly Arg Thr Gly Ile Ser Tyr Pro Gly Ala Leu
1 5 10 15
Leu Pro Gln Gly Phe Trp Ala Ala Val Glu Val Trp Leu Glu Arg
20 25 30
5/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
Pro Gln Val Ala Asn Lys Arg Leu Cys Gly Ala Arg Leu Glu Ala
'35 40 45
Arg Trp Ser Ala Ala Leu Pro Cys Ala Glu Ala Arg G1y Pro Gly
50 55 60
Thr Ser Ala Gly Ser Glu Gln Lys Glu Arg Gly Pro Gly Pro Gly
65 70 75
Gln Gly Ser Pro Gly Gly Gly Pro Gly Pro Arg Ser Leu Ser Gly
80 85 90
Pro Glu Gln Gly Thr Ala Cys Cys Glu Leu Glu Glu Ala Gln Gly
95 100 105
Gln Cys Gln Gln Glu Glu Ala Gln Arg Glu Ala Ala Ser Val Pro
110 115 120
Leu Arg Asp Ser Gly His Pro Gly His Ala Glu Gly Arg Glu Gly
125 130 135
Asp Phe Pro Ala Ala Asp Leu Asp Ser Leu Trp Glu Asp Phe Ser
140 145 150
Gln Ser Leu Ala Arg Gly Asn Ser Glu Leu Leu A1a Phe Leu Thr
155 160 165
Ser Ser G1y Ala Gly Ser G1n Pro Glu Ala Gln Arg Glu Leu Asp
170 175 180
Val Va1 Leu Arg Thr Val Ile Pro Lys Thr Ser Pro His Cys Pro
185 190 195
Leu Thr Thr Pro Arg Arg Glu Ile Val Val Gln Asp Val Leu Asn
200 205 210
Gly Thr I1e Thr Phe Leu Pro Leu Glu Glu Asp Asp Glu Gly Asn
215 220 225
Leu Lys Val Lys Met Ser Asn Val Tyr Gln Ile Gln Leu Ser His
230 235 240
Ser Lys Glu G1u Trp Phe Ile Ser Val Leu Ile Phe Cys Pro Glu
245 250 255
Arg Trp His Ser Asp Gly Ile Val Tyr Pro Lys Pro Thr Trp Leu
260 265 270
Gly Glu Glu Leu Leu Ala Lys Leu Ala Lys Trp Ser Val Glu Asn
275 280 285
Lys Lys Ser Asp Phe Lys Ser Thr Leu Ser Leu Ile Ser Ile Met
290 295 300
Lys Tyr Ser Lys Ala Tyr Gln Glu Leu Lys Glu Lys Tyr Lys Glu
305 310 315
Met Val Lys Val Trp Pro Glu Val Thr Asp Pro G1u Lys Phe Va1
320 325 330
Tyr Glu Asp Val Ala Ile Ala Ala Tyr Leu Leu Ile Leu Trp Glu
335 340 345
Glu Glu Arg Ala Glu Arg Gly Leu Thr Ala Arg Gln Ser Phe Val
350 355 360
Asp Leu G1y Cys Gly Asn Gly Leu Leu Val His Ile Leu Ser Ser
365 370 375
Glu Gly His Pro Gly Arg Gly Ile Asp Val Arg Arg Arg Lys Ile
380 385 390
Trp Asp Met Tyr Gly Pro Gln Thr Gln Leu Glu Glu Asp Ala Ile
395 400 405
Thr Pro Asn Asp Lys Thr Leu Phe Pro Asp Va1 Asp Trp Leu Ile
410 415 420
Gly Asn His Ser Asp Glu Leu Thr Pro Trp Ile Pro Val Ile Ala
425 430 435
Ala Arg Ser Ser Tyr Asn Cys Arg Phe Phe Val Leu Pro Cys Cys
440 445 450
Phe Phe Asp Phe Ile Gly Arg Tyr Ser Arg Arg Gln Ser Lys Lys
455 460 465
Thr Gln Tyr Arg Glu Tyr Leu Asp Phe Ile Lys Glu Val Gly Phe
470 475 480
Thr Cys Gly Phe His Val Asp Glu Asp Cys Leu Arg Ile Pro Ser
485 490 495
Thr Lys Arg Val Cys Leu Val Gly Lys Ser Arg Thr Tyr Pro Ser
6/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
500 505 510
Ser Arg Glu A1a Ser Va1 Asp Glu Lys Arg Thr Gln Tyr Ile Lys
515 520 525
Ser Arg Arg Gly Cys Pro Val Ser Pro Pro Gly Trp Glu Leu Ser
530 535 540
Pro Ser Pro Arg Trp Val Ala Ala G1y Ser Ala Gly His Cys Asp
545 550 555
Gly Gln Gln A1a Leu Asp Ala Arg Val Gly Cys Val Thr Arg Ala
560 565 570
Trp Ala Ala Glu His Gly Ala Gly Pro Gln Ala Glu Gly Pro Trp
575 580 585
Leu Pro Gly Phe His Pro Arg Glu Lys Ala Glu Arg Val Arg Asn
590 595 600
Cys Ala Ala Leu Pro Arg Asp Phe Ile Asp Gln Val Val Leu Gln
605 610 615
Val Ala Asn Leu Leu Leu Gly Gly Lys Gln Leu Asn Thr Arg Ser
620 625 630
Ser Arg Asn Gly Ser Leu Lys Thr Trp Asn Gly Gly Glu Ser Leu
635 640 645
Ser Leu Ala Glu Val Ala Asn Glu Leu Asp Thr Glu Thr Leu Arg
650 655 660
Arg Leu Lys Arg G1u Cys Gly Gly Leu Gln Thr Leu Leu Arg Asn
665 670 675
Ser His Gln Val Phe Gln Val Val Asn Gly Arg Val His Ile Arg
680 685 690
Asp Trp Arg Glu Glu Thr Leu Trp Lys Thr Lys G1n Pro Glu Ala
695 700 705
Lys Gln Arg Leu Leu Ser Glu Ala Cys Lys Thr Arg Leu Cys Trp
710 715 720
Phe Phe Met His His Pro Asp Gly Cys Ala Leu Ser Thr Asp Cys
725 730 735
Cys Pro Phe Ala His G1y Pro Ala Glu Leu Arg Pro Pro Arg Thr
740 745 750
Thr Pro Arg Lys Lys Ile Ser
755
<210> 7
<211> 1014
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 3846110CD1
<400> 7
Met Ile Met Gln Glu I1e Leu Thr Asp Asp Ile Pro Trp Lys Gly
1 5 10 15
Leu Tyr Gly Ser Val Val Lys Lys Ala Val Val Ser Gly Asn Tyr
20 25 30
Leu Glu Ala Asp Val Arg Leu Pro Lys Pro Tyr Tyr Asp Ile Val
35 40 45
Lys Ser Gly Ile His Val Lys His Lys Asp Arg Thr Met Asn Leu
50 55 60
Gln Asp Ile Arg Tyr Ile Leu Lys Asn Asp Leu Lys Asp Phe Thr
65 70 75
Gly A1a Gln Arg Thr Gln Pro Thr G1u Ser Pro Arg Val Gln Arg
80 85 90
Tyr Gly Leu His Pro Asp Val Asn Val Tyr Leu Gly Leu Thr Ser
95 100 105
G1u His Pro Arg Glu Thr Pro Asp Met Glu Ile Ile Glu Leu Lys
110 115 120
Glu Met Gly Ser Gln Pro His Ser Pro Arg Val His Ser Leu Phe
7/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
125 130 135
Thr Glu Gly Thr Leu Asp Pro Gln Ala Pro Asp Pro Cys Leu Met
140 145 150
Ala Arg G1u Thr Gln Asn Gln Asp Ala Pro Cys Pro Ala Pro Phe
155 160 165
Met Ala Glu Glu Ala Ser Ser Pro Ser Thr Gly Gln Pro Ser Leu
170 175 180
Cys Ser Phe Glu Ile Asn Glu I1e Tyr Ser Gly Cys Leu Ile Leu
185 190 195
G1u Asp Asp Ile Glu G1u Pro Pro Gly Ala Ala Ser Ser Leu Glu
200 205 210
Ala Asp Gly Pro Asn Gln Val Asp Glu Leu Lys Ser Met Glu Glu
215 220 225
Glu Leu Asp Lys Met Glu Arg Glu Ala Cys Cys Phe G1y Ser Glu
230 235 240
Asp Glu Ser Ser Ser Lys Ala Glu Thr Glu Tyr Ser Phe Asp Asp
245 250 255
Trp Asp Trp Gln Asn Gly Ser Leu Ser Ser Leu Ser Leu Pro Glu
260 ~ 265 270
Ser Thr Arg Glu A1a Lys Ser Asn Leu Asn Asn Met Ser Thr Thr
275 280 285
Glu Glu Tyr Leu Ile Ser Lys Cys Val Leu Asp Leu Lys I1e Met
290 295 300
Gln Thr Ile Met His Glu Asn Asp Asp Arg Leu Arg Asn Ile Glu
305 310 315
Gln Ile Leu Asp GlupVal Glu Met Lys Gln Lys Glu Gln Glu Glu
320 325 330
Arg Met Ser Leu Trp Ala Thr Ser Arg G1u Phe Thr Asn Ala Tyr
335 340 345
Lys Leu Pro Leu Ala Va1 G1y Pro Pro Ser Leu Asn Tyr Ile Pro
350 355 360
Pro Val Leu Gln Leu Ser Gly G1y Gln Lys Pro Asp Thr Ser Gly
365 370 375
Asn Tyr Pro Thr Leu Pro Arg Phe Pro Arg Met Leu Pro Thr Leu
380 385 390
Cys Asp Pro Gly Lys Gln Asn Thr Asp Glu Gln Phe Gln Cys Thr
395 400 405
Gln Gly Ala Lys Asp Ser Leu Glu Thr Ser Arg Ile Gln Asn Thr
410 415 420
Ser Ser Gln Gly Arg Pro Arg Glu Ser Thr Ala Gln A1a Lys Ala
425 430 435
Thr Gln Phe Asn Ser Ala Leu Phe Thr Leu Ser Ser His Arg Gln
440 445 450
Gly Pro Ser Ala Ser Pro Ser Cys His Trp Asp Ser Thr Arg Met
455 460 465
Ser Val Glu Pro Val Ser Ser Glu Ile Tyr Asn Ala Glu Ser Arg
470 475 480
Asn Lys Asp Asp Gly Lys Val His Leu Lys Trp Lys Met Glu Val
485 490 495
Lys Glu Met Ala Lys Lys Ala Ala Thr Gly Gln Leu Thr Val Pro
500 505 510
Pro Trp His Pro Gln Ser Ser Leu Thr Leu Glu Ser Glu Ala Glu
515 520 525
Asn Glu Pro Asp Ala Leu Leu G1n Pro Pro Ile Arg Ser Pro Glu
530 535 540
Asn Thr Asp Trp Gln Arg Val Ile Glu Tyr His Arg Glu Asn Asp
545 550 555
Glu Pro Arg Gly Asn Gly Lys Phe Asp Lys Thr Gly Asn Asn Asp
560 565 570
Cys Asp Ser Asp Gln His Gly Arg Gln Pro Arg Leu Gly Ser Phe
575 580 585
Thr Ser Ile Arg His Pro Ser Pro Arg Gln Lys Glu Gln Pro Glu
590 595 600
8/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
His Ser Glu Ala Phe Gln A1a Ser Ser Asp Thr Leu Val Ala Val
605 610 615
Glu Lys Ser Tyr Ser Thr Ser Ser Pro Ile Glu Glu Asp Phe Glu
620 625 630
Gly Ile Gln Gly Ala Phe Ala Gln Pro Gln Val Ser Gly Glu Glu
635 640 645
Lys Phe Gln Met Arg Lys Ile Leu Gly Lys Asn Ala Glu Ile Leu
6.50 655 660
Pro Arg Ser Gln Phe Gln Pro Val Arg Ser Thr Glu Asp Glu Gln
665 670 675
Glu G1u Thr Ser Lys Glu Ser Pro Lys Glu Leu Lys Glu Lys Asp
680 685 690
Ile Ser Leu Thr Asp Ile Gln Asp Leu Ser Ser Ile Ser Tyr Glu
695 700 705
Pro Asp Ser Ser Phe Lys Glu Ala Ser Cys Lys Thr Pro Lys Ile
710 715 720
Asn His Ala Pro Thr Ser Val Ser Thr Pro Leu Ser Pro Gly Ser
725 730 735
Val Ser Ser Ala Ala Ser G1n Tyr Lys Asp Cys Leu Glu Ser Ile
740 745 750
Thr Phe Gln Val Lys Thr Glu Phe Ala Ser Cys Trp Asn Ser Gln
755 760 765
G1u Phe Ile Gln Thr Leu Ser Asp Asp Phe I1e Ser Val Arg G1u
770 775 780
Arg Ala Lys Glu Leu Asp Ser Leu Leu Thr Ser Ser Glu Thr Pro
785 790 795
Pro Ser Arg Leu Thr Gly Leu Lys Arg Leu Ser Ser Phe Ile Gly
800 805 810
Ala Gly Ser Pro Ser Leu Val Lys Ala Cys Asp Ser Ser Pro Pro
815 820 825
His Ala Thr Gln Arg Arg Ser Leu Pro Lys Val Glu Ala Phe Ser
830 835 840
Gln His His Ile Asp Glu Leu Pro Pro Pro Ser Gln Glu Leu Leu
845 850 855
Asp Asp Ile Glu Leu Leu Lys Gln Gln Gln Gly Ser Ser Thr Val
860 865 870
Leu His Glu Asn Thr Ala Ser Asp Gly Gly Gly Thr Ala Asn Asp
875 880 885
Gln Arg His Leu Glu Glu Gln Glu Thr Asp Ser Lys Lys Glu Asp
890 895 900
Ser Ser Met Leu Leu Ser Lys Glu Thr Glu Asp Leu Gly Glu Asp
905 910 915
Thr Glu Arg Ala His Ser Thr Leu Asp G1u Asp Leu Glu Arg Trp
920 925 930
Leu Gln Pro Pro Glu Glu Ser Val G1u Leu Gln Asp Leu Pro Lys
935 940 945
Gly Ser Glu Arg Glu Thr Asn I1e Lys Asp Gln Lys Val Gly Glu
950 955 960
Glu Lys Arg Lys Arg Glu Asp Ser Ile Thr Pro Glu Arg Arg Lys
965 970 975
Ser Glu Gly Val Leu Gly Thr Ser Glu Glu Asp Glu Leu Lys Ser
980 985 990
Cys Phe Trp Lys Arg Leu Gly Trp Ser Glu Ser Ser Arg Ile Tle
995 1000 1005
Val Leu Asp Gln Ser Asp Leu Ser Asp
1010
<210> 8
<211> 342
<212> PRT
<213> Homo sapiens
<220>
9/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<221> misc_feature
<223> Incyte ID No: 1878279CD1
<400> 8
Met Met Cys Ser Arg Val Pro Ser Glu Gln Ser Ser Gly Thr Ser
1 5 10 15
Leu Leu Pro Lys Asp Gly Ala Pro Phe Ser Trp Asp Ser Leu Asp
20 25 , 30
Glu Asp Gly Leu Asp Asp Ser Leu Leu Glu Leu Ser Glu Gly Glu
35 40 45
Glu Asp Asp Gly Asp Val Asn Tyr Thr Glu Glu Glu Ile Asp Ala
50 55 60
Leu Leu Lys Glu Asp Asp Pro Ser Tyr Glu Gln Ser Ser Gly Glu
65 70 75
Asp Asp Gly Gly His Val G1u Lys Gly Glu Arg Gly Ser Gln Ile
80 85 , 90
Leu Leu Asp Thr Pro Arg Glu Lys Asn Ser Ser Tyr Ser Leu Gly
95 100 105
Pro Val A1a Glu Thr Pro Asp Leu Phe Lys Leu Pro G1n Leu Ser
110 115 120
Thr Ser Ser G1y His Gly Pro Ala His Thr Lys Pro Leu Asn Arg
125 130 135
Arg Ser Val Leu Glu Lys Asn Leu Ile Lys Val Thr Val Ala Pro
140 145 150
Phe Asn Pro Thr Val Cys Asp Ala Leu Leu Asp Lys Asp Glu Thr
155 160 165
Asp Ser Ser Lys Asp Thr Glu Lys Leu Ser Ser Leu Gly Glu Glu
170 175 180
Met Arg G1u Asp Gly Leu Ser Pro Asn Glu Ser Lys Leu Cys Thr
185 190 195
G1u Ser Glu Gly Ile Ser Pro Asn Asn Ser A1a Trp Asn Gly Pro
200 205 210
Gln Leu Ser Ser Ser Asn Asn Asn Phe Gln Gln Thr Va1 Ser Asp
215 220 225
Lys Asn Met Pro Asp Ser Glu Asn Pro Thr Ser Val Phe Ser Arg
230 235 240
Ile Ser Asp His Ser Glu Thr Pro Asn Met Glu Leu Ser Cys Arg
245 250 255
Asn Gly Gly Ser His Lys Ser Ser Cys Glu Met Arg Ser Leu Val
260 265 270
Va1 Ser Thr Ser Ser Asn Lys Gln Asp Val Leu Asn Lys Asp Ser
275 280 285
Gly Lys Met Lys Gly His Glu Arg Arg Leu Gly Lys Val Ile Pro
290 295 300
Val Leu Gln Thr Lys Thr Arg Thr Asn Val Pro Thr Phe Ser Gln
305 310 315
Ser Asn Leu Glu Gln Gln Lys Gln Leu Tyr Leu Arg Ser Val Ile
320 325 330
A1a His Ile Glu Asp Pro Glu Asp Thr Asn Gln Gly
335 340
<210> 9
<211> 415
<212> PRT
<213> Homo sapiens
<220>
<221> misc-feature
<223> Incyte ID No: 1848891CD1
<400> 9
Met Arg A1a Ala Asp Ser Gly Ser Trp Glu Arg Va1 Arg Gln Leu
1 5 10 15
10/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
Ala Ala Gln Gly Glu Pro Ala Pro Ser Cys G1y Ala Gly Ala Gly
20 25 30
Pro Ala Arg Pro Pro G1y Pro A1a Ala Cys Glu Gln Cys Val Asp
35 40 45
A1a Ala Gly Pro Gly Asp Arg Pro Arg Ala Gly Val Pro Arg Val
50 55 60
Arg Ala Asp Gly Asp Cys Ser Gln Pro Val Leu Leu Arg Glu Glu
65 70 75
Val Ser Arg Leu G1n Glu Glu Val His Leu Leu Arg Gln Met Lys
80 85 90
Glu Met Leu Ala Lys Asp Leu Glu Glu Ser G1n Gly Gly Lys Ser
95 100 105
Ser Glu Val Leu Ser Ala Thr Glu Leu Arg Val Gln Leu Ala G1n
110 115 120
Lys Glu G1n Glu Leu Ala Arg Ala Lys Glu Ala Leu Gln Ala Met
125 130 135
Lys A1a Asp Arg Lys Arg Leu Lys Gly Glu Lys Thr Asp Leu Val
140 145 150
Ser Gln Met Gln Gln Leu Tyr Ala Thr Leu Glu Ser Arg Glu Glu
155 160 165
Gln Leu Arg Asp Phe Ile Arg Asn Tyr Glu Gln His Arg Lys Glu
170 175 180
Ser Glu Asp Ala Va1 Lys Ala Leu Ala Lys Glu Lys Asp Leu Leu
185 190 195
Glu Arg Glu Lys Trp Glu Leu Arg Arg Gln Ala Lys Glu Ala Thr
200 205 210
Asp His A1a Thr Ala Leu Arg Ser Gln Leu Asp Leu Lys Asp Asn
215 220 225
Arg Met Lys Glu Leu Glu A1a Glu Leu Ala Met Ala Lys Gln Ser
230 235 240
Leu Ala Thr Leu Thr Lys Asp Val Pro Lys Arg His Ser Leu Ala
245 250 255
Met Pro G1y Glu Thr Va1 Leu Asn Gly Asn Gln Glu Trp Val Val
260 265 270
G1n Ala Asp Leu Pro Leu Thr A1a A1a Ile Arg Gln Ser G1n Gln
275 280 285
Thr Leu Tyr His Ser His Pro Pro His Pro Ala Asp Arg Gln Ala
290 295 300
Va1 Arg Val Ser Pro Cys His Ser Arg Gln Pro Ser Val Ile Ser
305 310 315
Asp Ala Ser Ala Ala Glu Gly Asp Arg Ser Ser Thr Pro Ser Asp
320 325 330
Ile Asn Ser Pro Arg His Arg Thr His Ser Leu Cys Asn Gly Asp
335 340 345
Ser Pro Gly Pro Val Gln Lys Asn Leu His Asn Pro Ile Val Gln
350 355 360
Ser Leu Glu Asp Leu Glu Asp Gln Lys Arg Lys Lys Lys Lys Glu
365 370 375
Lys Met Gly Phe Gly Ser Ile Ser Arg Val Phe Ala Arg Gly Lys
380 385 390
Gln Arg Lys Ser Leu Asp Pro Gly Leu Phe Asp Gly Thr Ala Pro
395 400 405
Asp Tyr Tyr Ile Glu Glu Asp Ala Asp Trp
410 415
<210> 10
<211> 665
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 2500251CD1
11/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<400> 10
Met A1a Gly Leu Ser Gly Ala Gln Ile Pro Asp Gly Glu Phe Thr
1 5 10 15
Ala Leu Val Tyr Arg Leu Ile Arg Asp Ala Arg Tyr Ala Glu Ala
20 25 30
Val Gln Leu Leu Gly Arg Glu Leu Gln Arg Ser Pro Arg Ser Arg
35 40 45
Ala Gly Leu Ser Leu Leu Gly Tyr Cys Tyr Tyr Arg Leu Gln Glu
50 55 60
Phe Ala Leu Ala Ala Glu Cys Tyr Glu Gln Leu Gly Gln Leu His
65 70 75
Pro Glu Leu Glu Gln Tyr Arg Leu Tyr Gln Ala Gln Ala Leu Tyr
80 85 90
Lys Ala Cys Leu Tyr Pro Glu Ala Thr Arg Val Ala Phe Leu Leu
95 100 105
Leu Asp Asn Pro Ala Tyr His Ser Arg Val Leu Arg Leu Gln Ala
110 115 120
Ala Ile Lys Tyr Ser Glu Gly Asp Leu Pro Gly Ser Arg Ser Leu
125 130 135
Val Glu Gln Leu Leu Ser Gly Glu Gly G1y Glu G1u Ser Gly Gly
140 145 150
Asp Asn Glu Thr Asp Gly Gln Val Asn Leu Gly Cys Leu Leu Tyr
155 160 165
Lys Glu Gly Gln Tyr Glu Ala Ala Cys Ser Lys Phe Ser Ala Thr
170 175 180
Leu Gln Ala Ser G1y Tyr Gln Pro Asp Leu Ser Tyr Asn Leu Ala
185 190 195
Leu Ala Tyr Tyr Ser Ser Arg Gln Tyr Ala Ser Ala Leu Lys His
200 205 210
Ile Ala Glu Ile Ile Glu Arg Gly Tle Arg Gln His Pro Glu Leu
215 220 225
Gly Va1 Gly Met Thr Thr Glu Gly Phe Asp Va1 Arg Ser Va1 Gly
230 235 240
Asn Thr Leu Val Leu His Gln Thr Ala Leu Val Glu Ala Phe Asn
245 250 255
Leu Lys Ala Ala Ile Glu Tyr Gln Leu Arg Asn Tyr Glu Val Ala
260 265 270
Gln Glu Thr Leu Thr Asp Met Pro Pro Arg Ala Glu Glu Glu Leu
275 280 285
Asp Pro Val Thr Leu His Asn Gln A1a Leu Met Asn Met Asp Ala
290 295 300
Arg Pro Thr Glu Gly Phe Glu Lys Leu G1n Phe Leu Leu G1n Gln
305 310 315
Asn Pro Phe Pro Pro G1u Thr Phe G1y Asn Leu Leu Leu Leu Tyr
320 325 330
Cys Lys Tyr G1u Tyr Phe Asp Leu Ala Ala Asp Val Leu Ala Glu
335 340 345
Asn Ala His Leu Thr Tyr Lys Phe Leu Thr Pro Tyr Leu Tyr Asp
350 355 360
Phe Leu Asp A1a Leu Ile Thr Cys Gln Thr Ala Pro G1u Glu Ala
365 370 375
Phe Ile Lys Leu Asp G1y Leu Ala Gly Met Leu Thr G1u Gln Leu
380 385 390
Arg Arg Leu Thr Lys G1n Val Gln Glu Ala Arg His Asn Arg Asp
395 400 405
Asp Glu Ala Ile Lys Lys Ala Val Asn Glu Tyr Asp Glu Thr Met
410 415 ' 420
Glu Lys Tyr Ile Pro Va1 Leu Met Ala Gln Ala Lys Ile Tyr Trp
425 430 435
Asn Leu Glu Asn Tyr Pro Met Val Glu Lys I1e Phe Arg Lys Ser
440 445 450
Va1 Glu Phe Cys Asn Asp His Asp Val Trp Lys Leu Asn Val Ala
455 460 465
12/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
His Val Leu Phe Met Gln G1u Asn Lys Tyr Lys Glu Ala Ile Gly
470 475 480
Phe Tyr Glu Pro Ile Va1 Lys Lys His Tyr Asp Asn Ile Leu Asn
485 490 495
Val Ser Ala Ile Val Leu Ala Asn Leu Cys Val Ser Tyr Tle Met
500 505 510
Thr Ser Gln Asn Glu Glu Ala Glu Glu Leu Met Arg Lys Ile Glu
515 520 525
Lys Glu Glu Glu Gln Leu Ser Tyr Asp Asp Pro Asn Arg Lys Met
530 535 540
Tyr His Leu Cys Ile Val Asn Leu Val Ile Gly Thr Leu Tyr Cys
545 550 555
Ala Lys G1y Asn Tyr Glu Phe Gly Ile Ser Arg Val Ile Lys Ser
560 565 570
Leu Glu Pro Tyr Asn Lys Lys Leu Gly Thr Asp Thr Trp Tyr Tyr
575 580 585
Ala Lys Arg Cys Phe Leu Ser Leu Leu Glu Asn Met Ser Lys His
590 595 600
Met Ile Val Ile His Asp Ser Val Tle G1n Glu Cys Val Gln Phe
605 610 615
Leu Gly His Cys Glu Leu Tyr Gly Thr Asn Ile Pro Ala Val Ile
620 625 630
Glu Gln Pro Leu Glu Glu Glu Arg Met His Val Gly Lys Asn Thr
635 640 645
Val Thr Asp Glu Ser Arg Gln Leu Lys Ala Leu Ile Tyr Glu Ile
650 655 660
Ile Gly Trp Asn Lys a
665
<210> 11
<211> 622
<212> PRT
<213> Homo sapiens
<220>
<221> misC_feature
<223> Incyte ID No: 55026561CD1
<400> 11
Met Asn His Phe Arg Lys Met Glu Val 21e Asn Leu Thr Thr Leu
1 5 10 15
Pro Met Ile Pro Val Asp G1u His Leu Ala Val Ser Leu Val Ala
20 25 30
Arg Asn Thr Met Val Lys Thr Val Arg Lys Glu Leu Glu Asn Asn
35 40 45
Pro Pro Ser Cys Leu Ile Gly Ser Met His Gln Val Asn Gln Lys
50 55 60
Ile Ala Asp Ile Asn Leu Arg Thr Glu Pro Ser Ala Asn Ser Leu
65 70 75
Ala Ile Glu Arg Phe Glu Leu Glu Lys Lys Ala Leu Arg Glu Lys
80 85 90
Thr Arg Ser Ser Pro Glu Asp Lys Val Lys Arg Gln Arg Lys Ser
95 100 105
Gln Tyr Ser Cys Lys G1y Ser Glu Leu Arg His Ala Arg Ser Ser
110 115 120
Val Ile Lys Arg Lys Thr Ala Asp Lys Asn Leu Leu Ala Glu Leu
125 130 135
Tyr G1n Tyr Ser Asn Phe Asn Ser Ser Lys Pro Asn Lys Leu Pro
140 145 150
Asn G1y Val Asp Phe Cys Asp Met Val Gly Asn Val Val Arg Ala
155 160 165
Glu Arg Asp Cys Leu Ser Gly Lys His Phe Cys Ser G1y Arg Glu
170 175 180
13/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
Leu Glu Lys Phe Leu Ser Ser Ser Ser Pro Arg Ala Ile Trp Leu
185 190 195
Asp Ser Phe Trp Trp Ile Phe His Glu Arg Tyr Gln Pro Asn Lys
200 205 210
Glu Leu Gln Asn Asn Leu Phe Asp Arg Ile Ala Gln His Tyr Ala
215 220 225
Leu Leu Leu Phe Arg Val Pro Lys Ser His Ser Glu Glu Ala Leu
230 235 240
Leu Lys Arg Leu Pro Ser Leu Leu Ser Lys Ala Val Tyr Thr Ser
245 250 255
Phe Cys Cys Cys Phe Pro Gln Ser Trp Phe Asp Thr His Glu Phe
260 265 270
Lys Ser Asp Ile Cys Asn Thr Met Ser Leu Trp Ile Ser Gly Thr
275 280 285
Tyr Pro Ser Pro Gln Ser Tyr Asp Ser Trp Asp Tyr Ser Glu Leu
290 295 300
Asp Pro Glu Arg Phe Arg Arg Glu Glu Leu Met Leu Tyr Arg Arg
305 310 315
Arg Leu Thr Lys Gly Arg Glu Phe Ser Leu Phe Ala Gly Lys Arg
320 325 330
Ala Phe Ser Gln Lys Pro Ala Gln Ser Arg Lys Phe Tyr His Pro
335 340 345
Gln Ser Ser Ser Ala Asn Ser Pro Ser Glu Lys Thr Ser Ser Ala
350 355 360
Lys G1n Asn Ser Glu Lys Ser Leu Arg Met Gln Asn Thr Ala Lys
365 370 375
Glu His His Cys G1n Thr Leu Va1 Leu Lys Lys Pro Thr G1n Glu
380 385 390
Val Lys Arg Ile Ser Glu Ala Arg Glu Cys Glu Asn Met Phe Pro
395 400 405
Lys Lys Ser Cys Ala Ala Cys Lys Ser Pro Glu Leu Thr Ser Asn
410 415 420
Leu Phe Asn Ile Tyr Gly Lys Ser Pro Leu Ile Val Tyr Phe Leu
425 430 435
G1n Asn Tyr Ala Ser Leu Gln Gln His Gly Lys Asn Val Leu Ile
440 445 450
Val Arg Arg Glu Lys Thr Thr Ser Thr Pro Asp Cys Thr Pro Thr
455 460 465
Tyr Thr Asp Val Ile Ser Glu Thr Leu Cys Ser Met Lys Lys Arg
470 475 480
Lys Asp Asn Leu Asn Gln Leu Tyr Gln His His Trp Thr Glu Trp
485 490 495
Asn Tyr Phe Asp Lys His Leu Lys Glu Leu G1n Asp Asn Phe Ser
500 505 510
Arg Glu Met Lys Asn Ile Gly Pro Lys Ala Ala Asp Thr Lys Lys
515 520 525
Ala Asn His Met Phe Ile Pro Pro Ser Ala Val Asn Glu Glu Ser
530 535 540
Pro Asp Lys Lys Thr Lys Gly Ser Leu Gln Arg Glu Ile Glu Phe
545 550 555
Lys Gly Cys Ser Asn Lys Asn His Gly Lys Gly Arg Ser Val Asn
560 565 570
Met Arg Gly Lys Glu Glu Arg Glu Arg.Glu Glu Lys Gln Lys Leu
575 580 585
Asn I1e Ser Phe His Ser Leu Pro Ser Pro Glu Glu Leu His Asn
590 595 600
Leu Glu Pro Gly Ser A1a Tyr Arg Ile Arg Asp Ile Ser Ala Thr
605 610 615
Arg Trp Ser Gly Thr Arg Lys
620
<210> 12
<211> 242
14190
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7502593CD1
<400> 12
Met Asp Asp Asp Asp Ala Lys Leu Lys Ala Glu Ile Glu Ala Glu
1 5 10 15
Leu Asp Lys Leu Ser Ile Ser Ser Leu Glu Lys Glu Asp Ile Glu
20 25 30
Ser Asp Ala Lys Ser Glu Thr Gln Ser Asp Asp Ser Asp Thr Asp
35 40 45
Ser Val Glu Leu Pro Glu Ser Val Leu His Cys Ile Asn Ile Ile
50 55 60
Lys Asn Arg Ser Lys A1a Val Glu Glu Leu Ile Leu Gln'Asp Leu
65 70 75
Glu Asp Thr Asp Ile Leu Ser Cys Ser Tyr Gly Ala Val Ser Asn
80 85 90
Asn His Met His Leu Arg Thr Gly Leu Ser Thr Glu Tyr G1u Glu
95 100 105
Ser Ser G1u Gln Leu Ile Lys Ile Leu Ser Glu Ile Glu Lys Glu
110 115 ' 120
Glu Phe Met Arg Ser Lys Thr Asp Cys Ala Thr Pro Asp Phe Val
125 130 135
Pro Glu Pro Ser Pro His Asp Leu Pro Met Asp Glu His Val Leu
140 145 150
Pro Asp Asp Ala Asp Ile Asn Phe G1y Tyr Cys Glu Val Glu Glu
155 160 165
Lys Cys Arg Gln Ser Phe Glu Ala Trp Gln Glu Lys Gln Lys Glu
170 175 180
Leu Glu Asp Lys Glu Lys Gln Thr Leu Lys Ala Gln Arg Asp Arg
185 190 195
G1u Glu Lys Gln Phe Gln Glu Lys Lys Lys Lys Arg His Cys Trp
200 205 210
Met Lys Gln Phe Lys Val Glu Lys Lys Lys Leu Glu Asn Ile Gln
215 220 225
Lys Val Phe Cys Phe Cys Phe Ser Cys Ile Phe Lys Ile Ser Ser
230 235 240
Tyr Leu
<210> 13
<211> 408
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7503957CD1
<400> 13
Met Met Cys Ser Arg Val Pro Ser Glu Gln Ser Ser Gly Thr Ser
1 5 10 15
Leu Leu Pro Lys Asp G1y Ala Pro Phe Ser Trp Asp Ser Leu Asp
20 25 30
Glu Asp Gly Leu Asp Asp Ser Leu Leu Glu Leu Ser Glu G1y Glu
35 40 45
Glu Asp Asp Gly Asp Val Asn Tyr Thr G1u Glu Glu Ile Asp Ala
50 55 60
Leu Leu Lys Glu Asp Asp Pro Ser Tyr Glu Gln Ser Ser Gly Glu
65 70 75
15/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
Asp Asp Gly Gly His Val Glu Lys Gly Glu Arg Gly Ser Gln Ile
80 85 90
Leu Leu Asp Thr Pro Arg Glu Lys Asn Ser Ser Tyr Ser Leu Gly
95 100 105
Pro Val Ala Glu Thr Pro Asp Leu Phe Lys Leu Pro G1n Leu Ser
110 115 120
Thr Ser Ser Gly His Gly Pro Ala His Thr Lys Pro Leu Asn Arg
125 130 135
Arg Ser Val Leu Glu Lys Asn Leu Ile Lys Val Thr Val Ala Pro
140 145 150
Phe Asn Pro Thr Val Cys Asp Ala Leu Leu Asp Lys Asp Glu Thr
155 160 165
Asp Ser Ser Lys Asp Thr Glu Lys Leu Ser Ser Leu Gly Glu Glu
170 175 180
Met Arg Glu Asp Gly Leu Ser Pro Asn Glu Ser Lys Leu Cys Thr
185 190 195
Glu Ser Glu Gly Ile Ser Pro Asn Asn Ser Ala Trp Asn Gly Pro
200 205 210
Gln Leu Ser Ser Ser Asn Asn Asn Phe Gln Gln Thr Val Ser Asp
215 220 225
Lys Asn Met Pro Asp Ser Glu Asn Pro Thr Ser Val Phe Ser Arg
230 235 240
Tle Ser Asp His Ser G1u Thr Pro Asn Met Glu Leu Ser Cys Arg
245 250 255
Asn Gly Gly Ser His Lys Ser Ser Cys Glu Met Arg Ser Leu Val
260 265 270
Val Ser Thr Ser Ser Asn Lys Gln Asp Val Leu Asn Lys Asp Ser
275 280 285
Gly Lys Met Lys Gly His Glu Arg Arg Leu Gly Lys Val Ile Pro
290 295 300
Val Leu Gln Thr Lys Thr Arg Thr Asn Va1 Pro Thr Phe Ser Gln
305 310 315
Ser Asn Leu G1u Gln Gln Lys Gln Leu Tyr Leu Arg Ser Val Ile
320 325 330
Ala His Ile Glu Asp Pro G1u Asp Thr Asn Gln Gly Tle Ser Gly
335 340 345
Glu Leu Cys Ala Leu Met Asp Gln Val His His Met G1n His Ser
350 355 360
Lys Trp Gln His Pro Ser Asp Leu Thr Thr Arg Asn Tyr Ala Arg
365 370 375
Arg Gln Lys His Leu Gln Arg Tyr Ser Leu Thr Gln Trp Val Asp
380 385 390
Arg Asn Met Arg Ser His His Arg Phe G1n Arg Leu Pro Asp Phe
395 400 405
Ser Tyr Ser
<210> 14
<211> 820
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7504415CD1
<400> 14
Met Thr Asp Thr Arg Arg Arg Val Lys Val Tyr Thr Leu Asn Glu
1 5 10 15
Asp Arg Gln Trp Asp Asp Arg Gly Thr G1y His Val Ser Ser Gly
20 25 30
Tyr Val Glu Arg Leu Lys Gly Met Ser Leu Leu Val Arg Ala Glu
35 40 45
16/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
Ser Asp Gly Ser Leu Leu Leu Glu'Ser Lys Ile Asn Pro Asn Thr
50 55 60
Ala Tyr Gln Lys Gln Gln Asp Thr Leu Ile Val Trp Ser G1u Ala
65 70 75
Glu Asn Tyr Asp Leu Ala Leu Ser Phe G1n Glu Lys Ala Gly Cys
80 85 90
Asp Glu Ile Trp Glu Lys Ile Cys Gln Val Gln Gly Lys Asp Pro
95 100 105
Ser Va1 Asp Ile Thr Gln Asp Leu Val Asp Glu Ser Glu Glu Glu
110 115 120
Arg Phe Asp Asp Met Ser Ser Pro Gly Leu Glu Leu Pro Ser Cys
125 130 135
Glu Leu Ser Arg Leu Glu Glu Ile A1a Glu Leu Val Ala Ser Ser
140 145 150
Leu Pro Ser Pro Leu Arg Arg G1u Lys Leu Ala Leu Ala Leu G1u
155 160 165
Asn Glu Gly Tyr Ile Lys Lys Leu Leu Glu Leu Phe His Val Cys
170 175 180
Glu Asp Leu Glu Asn I1e Glu Gly Leu His His Leu Tyr Glu Ile
185 190 195
Ile Lys Gly Ile Phe Leu Leu Asn Arg Thr Ala Leu Phe Glu Val
200 205 210
Met Phe Ser Glu Glu Cys Ile Met Asp Val Ile G1y Cys~Leu Glu
215 220 225
Tyr Asp Pro Ala Leu Ser Gln Pro Arg Lys His Arg Glu Phe Leu
230 235 240
Thr Lys Thr Ala Lys Phe Lys Glu Va1 Ile Pro Ile Ser Asp Pro
245 250 255
Glu Leu Lys Gln Lys Ile His Gln Thr Tyr Arg Val Gln Tyr Ile
260 265 270
Gln Asp Met Val Leu Pro Thr Pro Ser Val Phe Glu Glu Asn Met
275 280 285
Leu Ser Thr Leu His Ser Phe Ile Phe Phe Asn Lys Val G1u Ile
290 295 300
Va1 Gly Met Leu Gln G1u Asp Glu Lys Phe Leu Thr Asp Leu Phe
305 310 315
A1a Gln Leu Thr Asp Glu Ala Thr Asp Glu Glu Lys Arg Gln Glu
320 325 330
Leu Val Asn Phe Leu Lys Glu Phe Cys Ala Phe Ser Gln Thr Leu
335 340 345
Gln Pro Gln Asn Arg Asp Ala Phe Phe Lys Thr Leu Ser Asn Met
350 355 360
Gly I1e Leu Pro Ala Leu Glu Val Ile Leu G1y Met Asp Asp Thr
365 370 375
Gln Val Arg Ser A1a Ala Thr Asp Ile Phe Ser Tyr Leu Val Glu
380 385 390
Tyr Asn Pro Ser Met Val Arg Glu Phe Val Met Gln Glu Ala Gln
395 400 405
Gln Asn Asp Asp Asp Ile Leu Leu Ile Asn Leu I1e Ile Glu His
410 415 420
Met Ile Cys Asp Thr Asp Pro Glu Leu Gly Gly A1a Val Gln Leu
425 430 435
Met Gly Leu Leu Arg Thr Leu Val Asp Pro Glu Asn Met Leu Ala
440 445 450
Thr Ala Asn Lys Thr Glu Lys Thr Glu Phe Leu Gly Phe Phe Tyr
455 460 465
Lys His Cys Met His Val Leu Thr Ala Pro Leu Leu Ala Asn Thr
470 475 480
Thr Glu Asp Lys Pro Ser Lys Asp Asp Phe Gln Thr Ala Gln Leu
485 490 495
Leu Ala Leu Val Leu Glu Leu Leu Thr Phe Cys Val G1u His His
500 505 510
Thr Tyr His Ile Lys Asn Tyr Ile Ile Asn Lys Asp Ile Leu Arg
17/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
515 520 525
Arg Val Leu Val Leu Met Ala Ser Lys His A1a Phe Leu Ala Leu
530 535 540
Cys Ala Leu Arg Phe Lys Arg Lys Ile Ile Gly Leu Lys Asp Glu
545 550 555
Phe Tyr Asn Arg Tyr Ile Met Lys Ser Phe Leu Phe Glu Pro Val
560 ' 565 570
Val Lys Ala Phe Leu Asn Asn Gly Ser Arg Tyr Asn Leu Met Asn
575 580 585
Ser Ala Ile Ile Glu Met Phe Glu Phe Ile Arg Val Glu Asp Ile
590 595 600
Lys Ser Leu Thr Ala His Val Ile Glu Asn Tyr Trp Lys Ala Leu
605 610 615
Glu Asp Val Asp Tyr Val Gln Thr Phe Lys Gly Leu Lys Leu Arg
620 625 630
Phe Glu Gln Gln Arg Glu Arg Gln Asp Asn Pro Lys Leu Asp Ser
635 640 645
Met Arg Ser Ile Leu Arg Asn His Arg Tyr Arg Arg Asp Ala Arg
650 655 660
Thr Leu Glu Asp Glu Glu G1u Met Trp Phe Asn Thr Asp Glu Asp
665 670 675
Asp Met Glu Asp Gly Glu Ala Val Val Ser Pro Ser Asp Lys Thr
680 685 690
Lys Asn Asp Asp Asp Ile Met Asp Pro I1e Ser Lys Phe Met Glu
695 700 705
Arg Lys Lys Leu Lys Glu Ser Glu Glu Lys Glu Val Leu Leu Lys
710 715 720
Thr Asn Leu Ser Gly Arg Gln Ser Pro Ser Phe Lys Leu Ser Leu
725 730 735
Ser Ser Gly Thr Lys Thr Asn Leu Thr Ser Gln Ser Ser Thr Thr
740 745 750
Asn Leu Pro Gly Ser Pro Gly Ser Pro Gly Ser Pro Gly Ser Pro
755 760 765
Gly Ser Pro Gly Ser Val Pro Lys Asn Thr Ser Gln Thr Ala Ala
770 775 780
Ile Thr Thr Lys Gly Gly Leu Val G1y Leu Val Asp Tyr Pro Asp
785 790 795
Asp Asp Glu Asp Asp Asp Glu Asp Glu Asp Lys Glu Asp Thr Leu
800 805 810
Pro Leu Ser Lys Lys Ala Lys Phe Asp Ser
815 820
<210> 15
<211> 34
<212> PRT
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7504074CD1
<400> 15
Met Ser Val Pro Gly Pro Ser Ser Pro Asp Gly Ala Leu Thr Arg
1 5 10 15
Pro Pro Tyr Cys Leu Glu A1a Gly Glu Pro Thr Pro Asp Arg Thr
20 25 30
Lys Leu Tyr Cys
<210> 16
<211> 938
<212> PRT
<213> Homo sapiens
18/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<220>
<221> misc_feature
<223> Incyte ID No: 7502257CD1
<400> 16
Met Ala Glu Lys Arg Pro Leu Arg Thr Leu Gly Pro Val Met Tyr
1 5 10 15
Gly Lys Leu Pro Arg Leu Glu Thr Asp Ser Gly Leu Glu,His Ser
20 25 30
Leu Pro His Ser Val Gly Asn G1n Asp Pro Cys Thr Tyr Lys Gly
35 40 45
Ser Tyr Phe Ser Cys Pro Met Ala Gly Thr Pro Lys Ala Glu Ser
50 55 60
Glu Gln Leu Ala Ser Trp Thr Pro Tyr Pro Pro Leu Tyr Ser Thr
65 70 75
Gly Met A1a Gly Pro Pro Leu Gln Ala Asp Asn Leu Leu Thr Asn
80 85 90
Cys Leu Phe Tyr Arg Ser Pro Ala Glu Gly Pro Glu Lys Met Gln
95 100 105
Asp Ser Ser Pro Val G1u Leu Leu Pro Phe Ser Pro Gln Ala His
110 115 120
Ser Tyr Pro Gly Pro Pro Leu Ala Ala Pro Lys Pro Val Tyr Arg
125 130 135
Asn Pro Leu Cys Tyr Gly Leu Ser Thr Cys Leu Gly Glu Gly Ala
140 145 150
Val Lys Arg Pro Leu Asp Va1 Asp Trp Thr Leu Ala Thr Gly Pro
155 160 165
Leu Leu Pro Ser Ala Asp Pro Pro Cys Ser Leu Ala Pro Ala Pro
170 175 180
Ser Lys Gly Gln Thr Leu Asp G1y Thr Phe Leu Arg Gly Val Pro
185 190 195
Ala Glu Gly Ser Ser Lys Asp Ser Ser Gly Ser Phe Ser Pro Cys
200 205 210
Gln Pro Phe Leu Glu Lys Tyr Gln Thr Ile His Ser Thr Gly Phe
215 220 225
Leu Ala Ser Arg Tyr Thr Gly Pro Tyr Pro Arg Asn Ser Lys Gln
230 235 240
Ala Met Ser Glu Gly Pro Ser Ser Pro Trp Thr Gln Leu Ala Gln
245 250 255
Pro Leu Gly Pro Pro Cys Gln Asp Thr Gly Pro Thr His Tyr Pro
260 265 270
Pro Pro His His Pro Pro Pro His Pro Pro Gln Ala Leu Pro Cys
275 280 285
Pro Pro Ala Cys Arg His Pro Glu Lys Gln Gly Ser Tyr Ser Pro
290 295 300
Ala Leu Pro Leu Gln Pro Leu Gly Gly His Lys Gly Thr Gly Tyr
305 310 315
Gln Ala Gly Gly Leu Gly Ser Pro Tyr Leu Arg Gln Gln A1a Ala
320 325 330
Gln Ala Pro Tyr Ile Pro Pro Leu Gly Leu Asp Ala Tyr Pro Tyr
335 340 345
Pro Ser Ala Pro Leu Pro Ala Pro Ser Pro Gly Leu Lys Leu Glu
350 355 360
Pro Pro Leu Thr Pro Arg Cys Pro Leu Asp Phe Ala Pro Gln Thr
365 ' 370 375
Leu Ser Phe Pro Tyr A1a Arg Asp Asp Leu Ser Leu Tyr Gly Ala
380 385 390
Ser Pro Gly Leu Gly Gly Thr Pro Pro Ser Gln Asn Asn Val Arg
395 400 405
Ala Val Pro Gln Pro Gly Ala Phe Gln Arg Ala Cys Gln Pro Leu
410 415 420
Pro Ala Ser Gln Pro Cys Ser Glu Pro Val Arg Pro Ala Gln G1u
425 430 435
19/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
Ala Glu Glu Lys Thr Trp Leu Pro Ser Cys Arg Lys Glu Lys Leu
440 445 450
Gln Pro Arg Leu Ser Glu His Ser Gly Pro Pro Ile Val Ile Arg
455 460 465
Asp Ser Pro Val Pro Cys Thr Pro Pro Ala Leu Pro Pro Cys Ala
470 475 480
Arg Glu Cys Gln Ser Leu Pro Gln Lys Glu Asp Ala Arg Pro Pro
485 490 495
Ser Ser Pro Pro Met Pro Val Ile Asp Asn Val Phe Ser Leu Ala
500 505 510
Pro Tyr Arg Asp Tyr Leu Asp Val Pro Ala Pro Glu Ala Thr Thr
515 520 525
Glu Pro Asp Ser Ala Thr Ala Glu Pro Asp Ser Ala Pro Ala Thr
530 535 540
Ser Glu Gly Gln Asp Lys Gly Cys Arg Gly Thr Leu Pro Ala Gln
545 550 555
Glu Gly Pro Ser Gly Ser Lys Pro Leu Arg Gly Ser Leu Lys Glu
560 565 570
Glu Val A1a Leu Asp Leu Ser Val Arg Lys Pro Thr Ala Glu Ala
575 580 585
Ser Pro Val Lys Ala Ser Arg Ser Val Glu His Ala Lys Pro Thr
590 595 600
Ala Ala Met Asp Val Pro Asp Va1 Gly Asn Met Val Ser Asp Leu
605 610 615
Pro Gly Leu Lys Lys Ile Asp Thr Glu Ala Pro Gly Leu Pro Gly
620 625 630
Val Pro Val Thr Thr Asp Ala Met Pro Arg Thr Asn Phe His Ser
635 640 645
Ser Val Ala Phe Met Phe Arg Lys Phe Lys Ile Leu Arg Pro Ala
650 655 660
Pro Leu Pro Ala Ala Val Val Pro Ser Thr Pro Thr Ser Ala Pro
665 670 675
Ala Pro Thr G1n Pro Ala Pro Thr Pro Thr Ser Gly Pro Ile Gly
680 685 690
Leu Arg Ile Leu Ala Gln Gln Pro Leu Ser Val Thr Cys Phe Ser
695 700 705
Leu Ala Leu Pro Ser Pro Pro Ala Val Ala Val Ala Ser Pro Ala
710 715 720
Pro A1a Pro Ala Pro Ser Pro Ala Pro Ala Arg Ala Gln Ala Pro
725 730 735
Ala Ser Ala Arg Asp Pro Ala Pro Ala Pro Ala Pro Val Ala Gly
740 745 750
Pro Ala Pro Ala Ser Thr Ser Ala Pro Gly Asp Ser Leu Glu Gln
755 760 765
His Phe Thr G1y Leu His Ala Ser Leu Cys Asp Ala Ile Ser Gly
770 775 780
Ser Val Ala His Ser Pro Pro Glu Lys Leu Arg Glu Trp Leu Glu
785 790 795
Thr Ala G1y Pro Trp Gly Gln Ala Ala Trp G1n Asp Cys Gln Gly
800 805 810
Val Gln Gly Leu Leu Ala Lys Leu Leu Ser G1n Leu Gln Arg Phe
815 820 825
Asp Arg Thr His Arg Cys Pro Phe Pro His Val Val Arg Ala Gly
830 835 840
Ala Ile Phe Val Pro Ile His Leu Val Lys Glu Arg Leu Phe Pro
845 850 855
Arg Leu Pro Pro A1a Ser Val Asp His Val Leu Gln Glu His Arg
860 865 870
Val G1u Leu Arg Pro Thr Thr Leu Ser Glu Glu Arg Ala Leu'Arg
875 880 885
Glu Leu Ala Leu Pro Gly Cys Thr Ser Arg Met Leu Lys Leu Leu
890 895 900
Ala Leu Arg Gln Leu Pro Asp Ile Tyr Pro Asp Leu Leu Gly Leu
20190
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
905 910 915
Gln Trp Arg Asp Cys Val Arg Arg Gln Leu Gly Glu His Gly Ala
920 925 930
Ala Pro Val Ala Thr Gly Ala Val
935
<210> 17
<211> 253
<212> PRT
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 1315136CD1
<400> 17
Met Met Met Gly Cys Gly Glu Ser Glu Leu Lys Ser Ala Asp Gly
1 5 10 15
Glu Glu A1a Ala Ala Val Pro Gly Pro Pro Pro Glu Pro Gln Val
20 25 30
Pro Gln Leu Arg Ala Pro Val Pro Glu Pro Gly Leu Asp Leu Ser
35 40 45
Leu Ser Pro Arg Pro Asp Ser Pro Gln Pro Arg His Gly Ser Pro
50 55 60
Gly Arg Arg Lys Gly Arg A1a Glu Arg Arg Gly A1a Ala Arg Gln
65 70 75
Arg Arg Gln Val Arg Phe Arg Leu Thr Pro Pro Ser Pro Val Arg
80 85 90
Ser Glu Pro Gln Pro Ala Val Pro Gln Glu Leu Glu Met Pro Val
95 100 105
Leu Lys Ser Ser Leu Ala Leu Gly Leu Glu Leu Arg Ala Ala Ala
110 115 120
Gly Ser His Phe Asp Ala Ala Lys Ala Va1 Glu G1u Gln Leu Arg
125 130 135
Lys Ser Phe Gln Ile Arg Cys Gly Leu G1u Glu Ser Va1 Ser Glu
140 145 150
Gly Leu Asn Val Pro Arg Ser Lys Arg Leu Phe Arg Asp Leu Val
155 160 165
Ser Leu Gln Val Pro Glu Glu Gln Val Leu Asn Ala Ala Leu Arg
170 175 180
Glu Lys Leu Ala Leu Leu Pro Pro G1n Ala Arg Ala Pro His Pro
185 190 195
Lys Glu Pro Pro Gly Pro Gly Pro Asp Met Thr Ile Leu Cys Asp
200 205 210
Pro Glu Thr Leu Phe Tyr G1u Ser Pro His Leu Thr Leu Asp Gly
215 220 225
Leu Pro Pro Leu Arg Leu Gln Leu Arg Pro Arg Pro Ser Glu Asp
230 235 240
Thr Phe Leu Met His Arg Thr Leu Arg Arg Trp Glu A1a
245 250
<210> 18
<211> 723
<212> PRT
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 1379785CD1
<400> 18
Met Ala Glu Glu Glu Glu Thr Ala Ala Leu Thr Glu Lys Val Ile
1 5 10 15
21/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
Arg Thr Gln Arg Val Phe I1e Asn Leu Leu Asp Ser Tyr Ser Ser
20 25 30
Gly Asn Ile Gly Lys Phe Leu Ser Asn Cys Val Val Gly Ala Ser
35 40 45
Leu Glu Glu Ile Thr Glu Glu Glu Glu Glu Glu Asp Glu Asn Lys
50 55 60
Ser Ala Met Leu Glu Ala Ser Ser Thr Lys Ala Lys Glu G1y Thr
65 70 75
Phe Gln Ile Val Gly Thr Leu Ser Lys Pro Asp Ser Pro Arg Pro
80 85 90
Asp Phe Ala Val Glu Thr Tyr Ser Ala Ile Ser Arg Glu Asp Leu
95 100 105
Leu Met Arg Leu Leu Glu Cys Asp Val Ile Ile Tyr Asn Ile Thr
110 115 120
Glu Ser Ser Gln G1n Met Glu Glu Ala Ile Trp Ala Val Ser Ala
125 130 135
Leu Ser Glu Glu Val Ser His Phe Glu Lys Arg Lys Leu Phe Ile
140 145 150
Leu Leu Ser Thr Val Met Thr Trp Ala Arg Ser Lys Ala Leu Asp
155 160 165
Pro Glu Asp Ser Glu Val Pro Phe Thr Glu Glu Asp Tyr Arg Arg
170 175 180
Arg Lys Ser His Pro Asn Phe Leu Asp His Ile Asn Ala Glu Lys
185 190 195
Met Val Leu Lys Phe Gly Lys Lys Ala Arg Lys Phe Ala Ala Tyr
200 205 210
Val Va1 Ala Ala Gly Leu Gln Tyr Gly Ala Glu Gly Gly Met Leu
215 220 225
His Thr Phe Phe Lys Met Ala Trp Leu Gly Glu Ile Pro A1a Leu
230 235 240
Pro Val Phe Gly Asp G1y Thr Asn Val Ile Pro Thr Ile His Val
245 250 255
Leu Asp Leu Ala Gly Val Ile Gln Asn Val I1e Asp His Val Pro
260 265 270
Lys Pro His Tyr Leu Va1 Ala Val Asp Glu Ser Val His Thr Leu
275 280 285
Glu Asp Ile Val Lys Cys I1e Ser Lys Asn Thr Gly Pro G1y Lys
290 295 300
Ile Gln Lys Ile Pro Arg Glu Asn Ala Tyr Leu Thr Lys Asp Leu
305 310 315
Thr Gln Asp Cys Leu Asp His Leu Leu Val Asn Leu Arg Met Glu
320 325 330
Ala Leu Phe Val Lys Glu Asn Phe Asn Ile Arg Trp Ala Ala Gln
335 340 345
Thr Gly Phe Va1 G1u Asn Ile Asn Thr Ile Leu Lys Glu Tyr Lys
350 355 360
Gln Ser Arg Gly Leu Met Pro Ile Lys Ile Cys Ile Leu Gly Pro
365 370 375
Pro Ala Val Gly Lys Ser Ser Ile Ala Lys Glu Leu Ala Asn Tyr
380 385 390
Tyr Lys Leu His His Ile Gln Leu Lys Asp Val Ile Ser Glu Ala
395 400 405
Ile A1a Lys Leu Glu A1a Ile Val Ala Pro Asn Asp Val Gly Glu
410 415 420
Gly Glu G1u G1u Val Glu Glu Glu Glu Glu Glu Glu Asn Val Glu
425 430 435
Asp Ala Gln Glu Leu Leu Asp Gly I1e Lys Glu Ser Met Glu Gln
440 445 450
Asn Ala G1y Gln Leu Asp Asp Gln Tyr Ile Ile Arg Phe Met Lys
455 460 465
Glu Lys Leu Lys Ser Met Pro Cys Arg Asn Gln Gly Tyr Ile Leu
470 475 480
Asp Gly Phe Pro Lys Thr Tyr Asp Gln Ala Lys Asp Leu Phe Asn
22/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
485 490 495
Gln Glu Asp Glu Glu Glu Glu Asp Asp Val Arg Gly Arg Met Phe
500 505 510
Pro Phe Asp Lys Leu Ile Ile Pro Glu Phe Val Cys Ala Leu Asp
515 520 525
Ala Ser Asp Glu Phe Leu Lys G1u Arg Val Ile Asn Leu Pro Glu
530 535 540
Ser Ile Val Ala Gly Thr His Tyr Ser Gln Asp Arg Phe Leu Arg
545 550 555
Ala Leu Ser Asn Tyr Arg Asp Tle Asn Ile Asp Asp Glu Thr Val
560 565 570
Phe Asn Tyr Phe Asp Glu Leu Glu Ile His Pro Ile His Ile Asp
575 580 585
Val Gly Lys Leu Glu Asp Ala Gln Asn Arg Leu Ala Ile Lys Gln
590 595 600
Leu Ile Lys Glu Ile Gly Glu Pro Arg Asn Tyr Gly Leu Thr Asp
605 610 615
Glu Glu Lys Ala Glu Glu Glu Arg Lys Ala Ala Glu Glu Arg Leu
620 625 630
Ala Arg Glu Ala Ala Glu Glu Ala Glu Arg Glu His Gln G1u Ala
635 640 645
Va1 Glu Met Ala Glu Lys Ile Ala Arg Trp Glu Glu Trp Asn Lys
650 655 660
Arg Leu Glu Glu Val Lys Arg Glu Glu Arg Glu Leu Leu Glu Ala
665 670 675
G1n Ser Ile Pro Leu Arg Asn Tyr Leu Met Thr Tyr Val Met Pro
680 685 690
Thr Leu Ile Gln Gly Leu Asn Glu Cys Cys Asn Val Arg Pro Glu
695 700 705
Asp Pro Val Asp Phe Leu Ala Glu Tyr Leu Phe Lys Asn Asn Pro
710 715 720
Glu Ala Gln
<210> 19
<211> 253
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 2011166CD1
<400> 19
Met Ser Phe Ser Val His Asn Gln Lys Gly Ser Lys Arg Pro Leu
1 5 10 15
Pro Leu Glu Pro Leu Leu Phe Leu Gln Val Pro Arg Ser Asn Tyr
20 25 30
Leu His Phe Gln Glu G1u Lys Gln Arg Leu His Leu Lys Lys Phe
35 ' 40 45
Leu Leu Asp Arg Met Phe Leu Val A1a Lys Ile Gln Ala Asn Va1
50 55 60
Glu Arg Lys Asp Val A1a Asp Tyr Tyr Glu Gln Met Phe Gln Ser
65 70 75
Val Leu Lys His His Leu Gly Glu Ala Val Thr Gly Leu Leu Leu
80 85 90
Ile Tyr Pro Thr Ser Ile Leu His Ile Leu Glu Ser Ser Ser Asp
95 100 105
Thr Leu Tyr Lys Val Leu Leu Asp Tyr Ile Gly His'Val Lys Asp
110 115 120
Glu Thr Val Phe Phe Ile Gln Gln Met Lys Ile Ile Val Ile Ser
125 130 135
His Asn Ile Pro Met Arg Leu Phe Met Gln Trp His Val Ser Val
23/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
140 145 150
Ile Lys Val Pro Val Met Tyr Leu Asp Asp Val Thr Gln Ser Gln
155 160 165
Ser Leu Lys Glu Val Ile Thr Asp Phe Leu Thr Gln Thr His Lys
170 175 180
Leu Ser Leu Tyr Leu Cys Gln Thr Met Lys Val Gly Thr Lys Gly
185 190 195
Pro G1y Asp Asn Leu His Gln Val Ala Pro Asp Leu Leu Leu Pro
200 205 210
Glu Gln Ile Ile Lys Tyr Leu Cys Lys Ser Glu G1u Phe Met Asp
215 '220 225
Pro Ala Thr Phe Ile Asn Met Tyr Asn Arg Pro Ile His Ile Thr
230 235 240
Leu Asp Ser Glu Val Val Trp Pro Ala Pro Ser Arg Phe
245 250
<210> 20
<211> 154
<212> PRT
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 3434684CD1
<400> 20
Met Ser Val Gly Asn Ser Val Asn Asn Ser Pro Ala Ala Pro Gln
1 5 10 15
Ser Asp Phe Gln Leu Leu Pro Ala Gln Gly Ser Ser Leu Thr Asn
20 25 30
Phe Phe Pro Asp Val Gly Phe Asp Gln Gln Ser Met Arg Pro Gly
35 40 45
Pro Ala Phe Pro Gln Gln Val Pro Leu Val Gln Gln Gly Ser Arg
50 55 60
Glu Leu Gln Asp Ser Phe His Leu Arg Pro Ser Pro Tyr Ser Asn
65 70 75
Cys Gly Ser Leu Pro Asn Thr I1e Leu Pro G1u Asp Ser Ser Thr
80 85 ~ 90
Ser Leu Phe Lys Asp Leu Asn Ser Ala Leu Ala Gly Leu Pro Glu
95 100 105
Val Ser Leu Asn Val Asp Thr Pro Phe Pro Leu Glu Glu Glu Leu
110 115 120
Gln Ile G1u Pro Leu Ser Leu Asp Gly Leu Asn Met Leu Ser Asp
125 130 135
Ser Ser Met Gly Leu Leu Asp Pro Ser Val Glu Glu Thr Phe Arg
140 145 150
Ala Asp Arg Leu
<210> 21
<211> 566
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 5134056CD1
<400> 21
Met Arg Arg Gln Trp Gly Ser Ala Met Arg Ala Ala Glu Gln A1a
1 5 10 15
Gly Cys Met Val Ser Ala Ser Arg Ala Gly Gln Pro Glu Ala Gly
20 25 30
24/90
Ile Tyr Pro Thr Ser Ile Leu His Ile Leu Glu Ser Se
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
Pro Trp Ser Cys Ser Gly Val Ile Leu Ser Arg Ser Pro Gly Leu
35 40 45
Va1 Leu Cys His Gly Gly Ile Phe Val Pro Phe Leu Arg Ala Gly
50 55 60
Ser Glu Val Leu Thr Ala Ala Gly Ala Val Phe Leu Pro Gly Asp
65 70 75
Ser Cys Arg Asp Asp Leu Arg Leu His Val Gln Trp Ala Pro Thr
80 85 90
Ala Ala Gly Pro G1y Gly Gly Ala Glu Arg Gly Arg Pro Gly Leu
95 100 105
Cys Thr Pro Gln Cys Ala Ser Leu Glu Pro Gly Pro Pro Ala Pro
110 115 120
Ser Arg Gly Arg Pro Leu Gln Pro Arg Leu Pro A1a Glu Leu Leu
125 130 135
Leu Leu Leu Ser Cys Pro Ala Phe Trp Ala His Phe Ala Arg Leu
140 145 150
Phe Gly Asp Glu Ala Ala Glu Gln Trp Arg Phe Ser Ser A1a Ala
155 160 165
Arg Asp Asp Glu Val Ser G1u Asp Gly Glu Ala Asp Gln Leu Arg
170 175 180
A1a Leu Gly Trp Phe Ala Leu Leu Gly Val Arg Leu Gly Gln Glu
185 190 195
Glu Val G1u G1u Glu Arg G1y Pro Ala Met Ala Val Ser Pro Leu
200 205 210
Gly Ala Val Pro Lys Gly Ala Pro Leu Leu Val Cys Gly Ser Pro
215 220 225
Phe Gly Ala Phe Cys Pro Asp Ile Phe Leu Asn Thr Leu Ser Cys
230 235 240
Gly Val Leu Ser Asn Val Ala Gly Pro Leu Leu Leu Thr Asp Ala
245 250 255
Arg Cys Leu Pro Gly Thr Glu Gly Gly G1y Val Phe Thr Ala Arg
260 265 270
Pro A1a Gly Ala Leu Val A1a Leu Va1 Val Ala Pro Leu Cys Trp
275 280 285
Lys Ala Gly Glu Trp Val G1y Phe Thr Leu Leu Cys Ala Ala Ala
290 295 300
Pro Leu Phe Arg Ala Ala Arg Asp Ala Leu His Arg Leu Pro His
305 310 315
Ser Thr Ala Ala Leu A1a Ala Leu Leu Pro Pro Glu Val G1y Val
320 325 330
Pro Trp Gly Leu Pro Leu Arg Asp Ser Gly Pro Leu Trp Ala Ala
335 340 345
Ala Ala Val Leu Val Glu Cys Gly Thr Val Trp G1y Ser Gly Val
350 355 360
Ala Val Ala Pro Arg Leu Val Val Thr Cys Arg His Val Ser Pro
365 370 375
Arg Glu Ala Ala Arg Val Leu Val Arg Ser Thr Thr Pro Lys Ser
380 385 390
Val Ala Ile Trp Gly Arg Val Val Phe Ala Thr Gln Glu Thr Cys
395 400 405
Pro Tyr Asp I1e Ala Val Va1 Ser Leu Glu Glu Asp Leu Asp Asp
410 415 420
Val Pro I1e Pro Val Pro Ala Glu His Phe His Glu Gly Glu Ala
425 430 435
Val Ser Val Val Gly Phe Gly Val Phe Gly Gln Ser Cys Gly Pro
440 445 450
Ser Val Thr Ser Gly Ile Leu Ser Ala Val Val Gln Val Asn Gly
455 460 465
Thr Pro Val Met Leu Gln Thr Thr Cys Ala Val His Ser Gly Ser
470 475 480
Ser Gly Gly Pro Leu Phe Ser Asn His Ser Gly Asn Leu Leu Gly
485 490 495
Ile Ile Thr Ser Asn Thr Arg Asp Asn Asn Thr Gly Ala Thr Tyr
25/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
500 505 510
Pro His Leu Asn Phe Ser Tle Pro Ile Thr Val Leu Gln Pro Ala
515 520 525
Leu Gln Gln Tyr Ser Gln Thr Gln Asp Leu Gly Gly Leu Arg Glu
530 535 540
Leu Asp Arg Ala Ala Glu Pro Val Arg Val Val Trp Arg Leu Gln
545 550 555
Arg Pro Leu Ala Glu Ala Pro Arg Ser Lys Leu
560 565
<210> 22
<211> 234
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 5281724CD1
<400> 22
Met Glu Pro Ser Lys Thr Phe Met Arg Asn Leu Pro Ile Thr Pro
1 5 10 15
Gly Tyr Ser Gly Phe Val Pro Phe Leu Ser Cys Gln Gly Met Ser
20 25 30
Lys Glu Asp Asp Met Asn His Cys Val Lys Thr Phe Gln Glu Lys
35 40 45
Thr Gln Arg Tyr Lys Glu Gln Leu Arg Glu Leu Cys Cys Ala Val
50 55 60
Ala Thr Ala Pro Lys Leu Lys Pro Val Asn Ser Glu Glu Thr Val
65 70 75
Leu G1n Ala Leu His Gln Tyr Asn Leu Gln Tyr His Pro Leu Ile
80 85 90
Leu Glu Cys Lys Tyr Val Lys Lys Pro Leu Gln Glu Pro Pro Ile
95 100 105
Pro Gly Trp Ala Gly Tyr Leu Pro Arg Ala Lys Val Thr Glu Phe
110 115 120
Gly Cys Gly Thr Arg Tyr Thr Val Met Ala Lys Asn Cys Tyr Lys
125 130 135
Asp Phe Leu G1u I1e Thr Glu Arg Ala Lys Lys Ala His Leu Lys
140 145 150
Pro Tyr Glu Glu Ile Tyr Gly Va1 Ser Ser Thr Lys Thr Ser Ala
155 160 165
Pro Ser Pro Lys Val Leu Gln His Glu Glu Leu Leu Pro Lys Tyr
170 175 180
Pro Asp Phe Ser I1e Pro Asp Gly Ser Cys Pro Ala Leu Gly Arg
185 190 195
Pro Leu Arg Glu Asp Pro Lys Thr Pro Leu Thr Cys Gly Cys Ala
200 205 210
Gln Arg Pro Ser Ile Pro Cys Ser Gly Lys Met Tyr Leu Glu Pro
215 220 225
Leu Ser Ser Ala Lys Tyr Ala Glu Gly
230
<210> 23
<211> 268
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7502391CD1
<400> 23
26/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
Met Phe Val Glu Leu Asn Asn Leu Leu Asn Thr Thr Pro Asp Arg
1 5 10 15
Ala Glu Gln G1y Lys Leu Thr Leu Leu Cys Asp Ala Lys Thr Asp
20 25 30
Gly Ser Phe Leu Val His His Phe Leu Ser Phe Tyr Leu Lys Ala
35 40 45
Asn Cys Lys Val Cys Phe Va1 Ala Leu I1e Gln Ser Phe Ser His
50 55 60
Tyr Ser Ile Val Gly Gln Lys Leu Gly Val Ser Leu Thr Met Ala
65 70 75
Arg G1u Arg Gly G1n Leu Val Phe Leu Glu Gly Leu Lys Ser Ala
80 85 90
Val Asp Val Val Phe Gln Ala G1n Lys Glu Pro His Pro Leu Gln
95 100 105
Phe Leu Arg Glu Ala Asn Ala Gly Asn Leu Lys Pro Leu Phe Glu
110 115 120
Phe Val Arg Glu Ala Leu Lys Pro Val Asp Ser Gly Glu Ala Arg
125 130 135
Trp Thr Tyr Pro Va1 Leu Leu Val Asp Asp Leu Ser Val Leu Leu
140 145 150
Ser Leu Gly Met Gly Ala Val Ala Val Leu Asp Phe Ile His Tyr
155 160 165
Cys Arg Ala Thr Va1 Cys Trp Glu Leu Lys G1y Asn Met Val Val
170 175 180
Leu Val His Asp Ser Gly Asp Ala Glu Asp Glu Glu Asn Asp Ile
185 190 195
Leu Leu Asn Gly Leu Ser His G1n Ser His Leu Ile Leu Arg A1a
200 205 210
Glu Gly Leu Ala Thr Gly Phe Cys Arg Asp Val His Gly Gln Leu
215 220 225
Arg Ile Leu Trp Arg Arg Pro Ser Gln Pro Ala Val His Arg Asp
230 235 240
Gln Ser Phe Thr Tyr Gln Tyr Lys Ile Gln Asp Lys Arg Arg Val
245 250 255
Leu Phe Cys Gln Arg Asn Val Ser Cys Cys Ser Val Thr
260 265
<210> 24
<211> 694
<212> PRT
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte 2D No: 7502544CD1
<400> 24
Met Ala Thr Lys Thr Ala G1y Val Gly Arg Trp G1u Val Val Lys
1 5 10 15
Lys Gly Arg Arg Pro Gly Val Gly Ala Gly A1a Gly Gly Arg Gly
20 25 30
Gly Gly Arg Asn Arg Arg Ala Leu G1y Glu Ala Asn Gly Val Trp
35 40 45
Lys Tyr Asp Leu Thr Pro Ala Ile Gln Thr Thr Ser Thr Leu Tyr
50 55 60
Glu Arg Gly Phe Glu Asn Ile Met Lys Arg Gln Asn Lys Glu Gln
65 70 75
Val Pro Pro Pro Ala Val G1u Pro Lys Lys Pro Gly Asn Lys Lys
80 85 90
Gln Pro Lys Lys Val Ala Thr Pro Pro Asn Gln Asn Gln Lys Gln
95 100 105
Gly Arg Phe Arg Ser Leu Glu Glu Ala Leu Lys Ala Leu Asp Val
110 115 120
27/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
Ala Asp Leu Gln Lys Glu Leu Asp Lys Ser Gln Ser Val Phe Ser
125 130 135
Gly Asn Pro Ser I1e Trp Leu Lys Asp Leu Ala Ser Tyr Leu Asn
140 145 150
Tyr Lys Leu Gln Ala Pro Leu Ser Glu Pro Thr Leu Ser Gln His
155 160 165
Thr His Gly Leu Trp A1a Leu Asp Tyr Pro Tyr Ser Leu Val Ser
170 175 180
Arg Glu Leu Arg Gly Ile I1e Arg Gly Leu Leu Ala Lys Ala Ala
185 190 195
G1y Ser Leu Glu Leu Phe Phe Asp His Cys Leu Phe Thr Met Leu
200 205 210
Gln Glu Leu Asp Lys Thr Pro Gly Glu Ser Leu His Gly Tyr Arg
215 220 225
Ile Cys Ile Gln Ala Tle Leu Gln Asp Lys Pro Lys Ile Ala Thr
230 235 240
Ala Asn Leu Gly Lys Phe Leu Glu Leu Leu Arg Ser His Gln Ser
245 250 255
Arg Pro Ala Lys Cys Leu Thr Ile Met Trp Ala Leu Gly Gln Ala
260 265 270
Gly Phe Ala Asn Leu Thr Glu Gly Leu Lys Val Trp Leu Gly Ile
275 280 285
Met Leu Pro Val Leu Gly Ile Lys. Ser Leu Ser Pro Phe Ala Tle
290 295 300
Thr Tyr Leu Asp Arg Leu Leu Leu Met His Pro Asn Leu Thr Lys
305 310 315
Gly Phe Gly Met Ile Gly Pro Lys Asp Phe Phe Pro Leu Leu Asp
320 325 330
Phe Ala Tyr Met Pro Asn Asn Ser Leu Thr Pro Ser Leu Gln Glu
335 340 345
Gln Leu Cys Gln Leu Tyr Pro Arg Leu Lys Met Leu A1a Phe Gly
350 355 360
A1a Lys Pro Asp Ser Thr Leu His Thr Tyr Phe Pro Ser Phe Leu
365 370 375
Ser Arg A1a Thr Pro Ser Cys Pro Pro Glu Met Lys Lys Glu Leu
380 385 390
Leu Ser Ser Leu Thr Glu Cys Leu Thr Val Asp Pro Leu Ser Ala
395 400 405
Ser Val Trp Arg Gln Leu Tyr Pro Lys His Leu Ser Gln Ser Ser
410 415 420
Leu Leu Leu Glu His Leu Leu Ser Ser Trp Glu Gln Ile Pro Lys
425 430 435
Lys Val Gln Lys Ser Leu Gln Glu Thr Ile Gln Ser Leu Lys Leu
440 445 450
Thr Asn Gln Glu Leu Leu Arg Lys G1y Ser Ser Asn Asn G1n Asp
455 460 465
Val Val Thr Cys Asp Met Ala Cys Lys Gly Leu Leu Gln Gln Val
470 475 480
Gln Gly Pro Arg Leu Pro Trp Thr Arg Leu Leu Leu Leu Leu Leu
485 490 495
Val Phe Ala Val Gly Phe Leu Cys His Asp Leu Arg Ser His Ser
500 505 510
Ser Phe Gln Ala Ser Leu Thr Gly Arg Leu Leu Arg Ser Ser Gly
515 520 525
Phe L~u Pro Ala Ser Gln Gln Ala Cys Ala Lys Leu Tyr Ser Tyr
530 535 540
Ser Leu Gln Gly Tyr Ser Trp Leu Gly Glu Thr Leu Pro Leu Trp
545 550 555
Gly Ser His Leu Leu Thr Va1 Val Arg Pro Ser Leu Gln Leu Ala
560 565 570
Trp Ala His Thr Asn Ala Thr Val Ser Phe Leu Ser Ala His Cys
575 580 585
Ala Ser His Leu Ala Trp Phe G1y Asp Ser Leu Thr Ser Leu Ser
28/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
590 595 G00
Gln Arg Leu Gln Ile G1n Leu Pro Asp Ser Va1 Asn G1n Leu Leu
605 610 615
Arg Tyr Leu Arg Glu Leu Pro Leu Leu Phe His Gln Asn Va1 Leu
620 625 630
Leu Pro Leu Trp His Leu Leu Leu Glu Ala Leu Ala Trp Ala Gln
635 640 645
Glu His Cys His Glu Ala Cys Arg Gly Glu Val Thr Trp Asp Cys
650 655 660
Met Lys Thr Gln Leu Ser Glu Ala Val His Trp Thr Trp Leu Cys
665 670 675
Leu Gln Asp Ile Thr Val Ala Phe Leu Asp Trp Ala Leu Ala Leu
680 685 690
Ile Ser Gln Gln
<210> 25
<211> 519
<212> PRT
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 2858465CD1
<400> 25
Met Ala Pro Ile Pro Lys Thr Val Gly Arg Tle Lys Leu Asp Cys
1 5 10 15
Ser Leu Arg Pro Ser Cys Pro Leu Glu Val Ala Ala Ala Pro Lys
20 25 30
Leu Cys Lys Glu Phe Gly Pro Glu Asp Tyr Gly Glu Glu Asp Ile
35 40 45
Val Asp Phe Leu Arg Arg Leu Va1 G1u Ser Asp Pro Gln Gly Leu
50 55 60
His Arg Ile His Val Asp Gly Ser Ser Gly Arg Leu Gln Leu Trp
65 70 75
His His Asp Tyr Leu Leu G1y His Leu Asp Asp Glu Gly Lys Ser
80 85 90
Thr Gly Gln Ser Asp Arg Gly Lys Gly Ala Glu Gly Leu Gly Thr
95 100 105
Tyr Cys Gly Leu Arg Lys Ser Phe Leu Tyr Pro Pro Gln Glu Ser
110 115 120
Glu Pro Cys Pro Gln Ser Pro Ser Ala Ser A1a Thr Phe Pro Ser
125 130 135
Val Ser Asp Ser Leu Leu Gln Val Ala Met Pro Gln Lys Leu Leu
140 145 150
Val Thr Glu Glu Glu A1a Asn Arg Leu Ala Glu G1u Leu Va1 Ala
155 160 165
Glu Glu G1u Arg Met Lys G1n Lys Ala Glu Lys Lys Arg Leu Lys
170 175 180
Lys Lys Arg Gln Lys Glu Arg Lys Arg Gln Glu Arg Leu Glu Gln
185 190 195
Tyr Cys Gly Glu Pro Lys Ala Ser Thr Thr Ser Asp Gly Asp Glu
200 205 210
Ser Pro Pro Ser Ser Pro Gly Asn Pro Val G1n Gly Gln Cys Gly
215 220 225
Glu Glu Glu Asp Ser Leu Asp Leu Ser Ser Thr Phe Val Ser Leu
230 235 240
Ala Leu Arg Lys Val Gly Asp Trp Pro Leu Ser Ala Arg Arg Glu
245 250 255
Lys Gly Leu Asn Gln Glu Pro Gln Gly Arg Gly Leu Ala Leu Gln
260 265 270
Lys Met Gly Gln Glu Glu Glu Ser Pro Pro Arg Glu Glu Arg Pro
29/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
275 280 285
Gln Gln Ser Pro Lys Val Gln Ala Ser Pro Gly Leu Leu A1a Ala
290 295 300
Ala Leu Gln Gln Ser Gln Glu Leu Ala Lys Leu Gly Thr Ser Phe
305 310 315
Ala Gln Asn Gly Phe Tyr His Glu Ala Val Val Leu Phe Thr Gln
320 325 330
Ala Leu Lys Leu Asn Pro Gln Asp His Arg Leu Phe Gly Asn Arg
335 340 345
Ser Phe Cys His Glu Arg Leu G1y Gln Pro A1a Trp Ala Leu Ala
350 355 360
Asp Ala Gln Val Ala Leu Thr Leu Arg Pro Gly Trp Pro Arg Gly
365 370 375
Leu Phe Arg Leu Gly Lys Ala Leu Met Gly Leu Gln Arg Phe Arg
380 385 390
Glu Ala Ala A1a Val Phe Gln Glu Thr Leu Arg Gly Gly Ser Gln
395 400 405
Pro Asp Ala Ala Arg Glu Leu Arg Ser Cys Leu Leu His Leu Thr
410 415 420
Leu Gln Gly Gln Arg G1y Gly Ile Cys Ala Pro Pro Leu Ser Pro
425 430 435
Gly Ala Leu Gln Pro Leu Pro His Ala Glu Leu Ala Pro Ser Gly
440 445 450
Leu Pro Ser Leu Arg Cys Pro Arg Ser Thr Ala Leu Arg Ser Pro
455 460 465
Gly Leu Ser Pro Leu Leu His Tyr Pro Ser Cys His Arg Ser His
470 475 480
Pro Asn Gln Pro Leu Ser Arg Thr Gln Ser Arg Arg Pro His Pro
485 490 495
Leu Lys Pro Gln Asp Pro Ser Lys Gly Trp Asp Ile Leu Gly Leu
500 505 510
G1y Leu Gln His Leu Ser Gln Ala Arg
515
<210> 26
<211> 216
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7503455CD1
<400> 26
Met Ala Leu Asn Lys Asn His Ser Glu G1y Gly Gly Va1 Ile Val
1 5 10 15
Asn Asn Thr Glu Ser Ile Leu Met Ser Tyr Asp His Val Glu Leu
20 25 30
Thr Phe Asn Asp Met Lys Asn Val Pro Glu Ala Phe Lys Gly Thr
35 40 45
Lys Lys G1y Thr Va1 Tyr Leu Thr Pro Tyr Arg Val Ile Phe Leu
50 55 60
Ser Lys Gly Lys Asp Ala Met Gln Ser Phe Met Met Pro Phe Tyr
65 70 75
Leu Met Lys Asp Cys Glu Ile Lys Gln Pro Val Phe Gly Ala Asn
80 85 90
Tyr Ile Lys Gly Thr Val Lys Ala Glu Ala Gly Gly Gly Trp Glu
95 100 105
Gly Ser Ala Ser Tyr Lys Leu Thr Phe Thr A1a Gly Gly A1a Ile
110 115 120
Glu Phe Gly Gln Arg Met Leu Gln Va1 Ala Ser Gln Glu Phe Tyr
125 130 135
Pro Gly Pro Pro Met Met Asp Gly Ala Met Gly Tyr Val Gln Pro
30/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
140 145 150
Pro Pro Pro Pro Tyr Pro Gly Pro Met Glu Pro Pro Val Ser Gly
155 160 165
Pro Asp Val Pro Ser Thr Pro Ala Ala Glu Ala Lys A1a Ala Glu
170 175 180
Ala Ala Ala Ser A1a Tyr Tyr Asn Pro Gly Asn Pro His Asn Val
185 190 195
Tyr Met Pro Thr Ser Gln Pro Pro Pro Pro Pro Tyr Tyr Pro Pro
200 205 210
Glu Asp Lys Lys Thr Gln
215
<210> 27
<211> 110
<212> PRT
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7503479CD1
<400> 27
Met Ala Val Cys Ile Ala Val Ile Ala Lys Glu Asn Tyr Pro Leu
1 5 10 15
Tyr Ile Arg Ser Thr Pro Thr Glu Asn Glu Leu Lys Phe His Tyr
20 25 30
Met Val His Thr Ser Leu Asp Val Val Asp Glu Lys Ile Ser Ala
35 40 45
Met Gly Lys Ala Leu Val Asp Gln Arg Glu Leu Tyr Leu Gly Leu
50 55 60
Leu Tyr Pro Thr G1u Asp Tyr Lys Met Phe Arg Lys Leu His Asn
65 70 75
Ser Tyr Thr Asp Val Met Cys Asn Pro Phe Tyr Asn Pro Gly Asp
80 85 90
Arg Ile Gln Ser Ser Arg Ala Phe Asp Asn Met Val Thr Ser Met
95 100 105
Met Ile Gln Val Cys
110
<210> 28
<211> 642
<212> PRT
<213> Homo sapiens
<220>
<221> misC_feature
<223> Incyte ID No: 7218127CD1
<400> 28
Met Gly Val Asp Ser Arg Thr Ser Cys Ser Pro Gln Lys Ala Gln
1 5 10 15
Glu Ala Asn Lys Ala Arg Pro Ser Ala Trp Glu Pro Ala Ala Gly
20 25 30
Asn Ser Pro Ala Arg Ala Ser Val Pro Ala Ala Pro Asn Pro Ala
35 40 45
Ala Thr Ser Ala Thr Ser Val His Val Arg Ser Pro Ala Arg Pro
50 55 60
Ser Glu Ser Arg Leu Ala Pro Thr Pro Thr Glu Gly Lys Val Arg
65 70 75
Pro Arg Val Thr Asn Ser Ser Pro Met Gly Trp Ser Ser Ala Ala
80 85 90
Pro Cys Thr Ala Ala Ala Ala Ser His Pro Ala Val Pro Pro Ser
95 100 105
31/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
Ala Pro Asp Pro Arg Pro Ala Thr Pro Gln Gly Gly Gly A1a Pro
110 115 120
Arg Val Ala Ala Pro Gln Thr Thr Leu Ser Ser Ser Ser Thr Ser
125 130 135
Ala Ala Thr Val Asp Pro Pro Ala Trp Thr Pro Ser Ala Ser Arg
140 145 150
Thr Gln Gln A1a Arg Asn Lys Phe Phe Gln Thr Ser Ala Val Pro
155 160 165
Pro Gly Thr Ser Leu Ser Gly Arg Gly Pro Thr Pro Ser Leu Val
170 175 180
Leu Ser Lys Asp Ser Ser Lys Glu Gln Ala Arg Asn Phe Leu Lys
185 190 195
Gln Ala Leu Ser Ala Leu Glu Glu Ala Gly Ala Pro Ala Pro Gly
200 205 210
Arg Pro Ser Pro Ala Thr Ala Ala Val Pro Ser Ser Gln Pro Lys
215 220 225
Thr Glu Ala Pro G1n Ala Ser Pro Leu Ala Lys Pro Leu Gln Ser
230 235 240
Ser Ser Pro Arg Val Leu Gly Leu Pro Ser Arg Met Glu Pro Pro
245 250 255
Ala Pro Leu Ser Thr Ser Ser Thr Ser Gln Ala Ser Ala Leu Pro
260 265 270
Pro Ala Gly Arg Arg Asn Leu Ala Glu Ser Ser Gly Val Gly Arg
275 280 285
Val Gly Ala Gly Ser Arg Pro Lys Pro Glu A1a Pro Met Ala Lys
290 295 300
Gly Lys Ser Thr Thr Leu Thr Gln Asp Met Ser Thr Ser Leu Gln
305 310 315
Glu Gly Gln Glu Asp Gly Pro Ala Gly Trp Arg Ala Asn Leu Lys
320 325 330
Pro Val Asp Arg Arg Ser Pro Ala Glu Arg Thr Leu Lys Pro Lys
335 340 345
Glu Pro Arg Ala Leu Ala Glu Pro Arg Ala Gly Glu A1a Pro Arg
350 355 360
Lys Val Ser Gly Ser Phe Ala Gly Ser Val His Ile Thr Leu Thr
365 370 375
Pro Val Arg Pro Asp Arg Thr Pro Arg Pro Ala Ser Pro Gly Pro
380 385 390
Ser Leu Pro Ala Arg Ser Pro Ser Pro Pro Arg Arg Arg Arg Leu
395 400 405
Ala Val Pro Ala Ser Leu Asp Val Cys Asp Asn Trp Leu Arg Pro
410 415 420
Glu Pro Pro Gly Gln Glu Ala Arg Va1 Gln Ser Trp Lys Glu Glu
425 430 435
Glu Lys Lys Pro His Leu Gln Gly Lys Pro Gly Arg Pro Leu Ser
440 445 450
Pro Ala Asn Val Pro Ala Leu Pro Gly Glu Thr Val Thr Ser Pro
455 460 465
Val Arg Leu His Pro Asp Tyr Leu Ser Pro Glu Glu Ile Gln Arg
470 475 480
Gln Leu Gln Asp Ile Glu Arg Arg Leu Asp Ala Leu Glu Leu Arg
485 490 495
G1y Val Glu Leu G1u Lys Arg Leu Arg Ala Ala Glu Gly Asp Asp
500 505 510
Ala Glu Asp Ser Leu Met Val Asp Trp Phe Trp Leu Ile His G1u
515 520 525
Lys Gln Leu Leu Leu Arg Gln Glu Ser Glu Leu Met Tyr Lys Ser
530 535 540
Lys Ala G1n Arg Leu G1u Glu Gln Gln Leu Asp Ile Glu Gly Glu
545 550 555
Leu Arg Arg Leu Met Ala Lys Pro Glu Ala Leu Lys Ser Leu G1n
560 565 570
Glu Arg Arg Arg Glu Gln Glu Leu Leu Glu Gln Tyr Val Ser Thr
32/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
575 580 585
Val Asn Asp Arg Ser Asp Ile Val Asp Ser Leu Asp Glu Asp Arg
590 595 600
Leu Arg Glu G1n Glu Glu Asp Gln Met Leu Arg Asp Met Ile Glu
605 610 615
Lys Leu Gly Leu Gln Arg Lys Lys Ser Lys Phe Arg Leu Ser Lys
620 625 630
Ile Trp Ser Pro Lys Ser Lys Ser Ser Pro Ser Gln
635 640
<210> 29
<211> 489
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 1688943CD1
<400> 29
Met Asn Lys Leu Ser Pro Val Leu Leu Phe Leu Asn Gln Gln Asn
1 5 10 15
Tyr Gln Ile Asp Lys Asp Val Glu Asp Lys Arg Gln Lys Ala Ile
20 25 30
Glu Glu Phe Phe Thr Lys Asp Val Ile Val Pro Ser Pro Trp Thr
35 40 45
Asp His G1u Gly Lys Gln Leu Ser Gln Cys His Ser Ser Lys Cys
50 55 60
Thr Asn Ile Asn Ser Asp Ser Pro Val Gly Lys Lys Leu Thr Ile
65 70 75
His Ser Glu Lys Ser Asp Ala Ala Cys Gln Thr Leu Leu Ser Leu
80 85 90
Pro Val Asp Phe Asn Leu Glu Asn Ile Leu Gly Asp Tyr Phe Arg
95 100 105
Ala Asp Glu Phe Ala Asp Gln Ser Pro G1y Asn Leu Ser Ser Ser
110 115 120
Ser Leu Arg Arg Lys Leu Phe Leu Asp Gly Asn G1y Ser Ile Ser
125 130 135
Asp Ser Leu Pro Ser Ala Ser Pro Gly Ser Pro His Ser Gly Val
140 145 150
Gln Thr Ser Leu Glu Met Phe Tyr Ser Ile Asp Leu Ser Pro Va1
155 160 165
Lys Cys Arg Ser Pro Leu Gln Thr Pro Ser Ser Gly Gln Phe Ser
170 175 180
Ser Ser Pro Ile Gln Ala Ser Ala Lys Lys Tyr Ser Leu Gly Ser
185 190 195
Ile Thr Ser Pro Ser Pro Ile Ser Ser Pro Thr Phe Ser Pro Ile
200 205 210
Glu Phe Gln Ile Gly Glu Thr Pro Leu Ser Glu G1n Arg Lys Phe
215 220 225
Thr Val His Ser Pro Asp Ala Ser Ser Gly Thr Asn Ser Asn Gly
230 235 240
Ile Thr Asn Pro Cys Ile Arg Ser Pro Tyr Ile Asp Gly Cys Ser
245 250 255
Pro Ile Lys Asn Trp Ser Pro Met Arg Leu Gln Met Tyr Ser Gly
260 265 270
Gly Thr Gln Tyr Arg Thr Ser Val Ile Gln Ile Pro Phe Thr Leu
275 280 285
Glu Thr G1n Gly Glu Asp G1u G1u Asp Lys G1u Asn Ile Pro Ser
290 295 300
Thr Asp Val Ser Ser Pro Ala Met Asp Ala Ala Gly Ile His Leu
305 310 315
Arg Gln Phe Ser Asn Glu Ala Ser Thr His Gly Thr His Leu Va1
33/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
320 325 330
Val Thr Ala Met Ser Val Thr Gln Asn Gln Ser Ser A1a Ser Glu
335 340 345
Lys Glu Leu Ala Leu Leu G1n Asp Val Glu Arg Glu Lys Asp Asn
350 355 360
Asn Thr Val Asp Met Val Asp Pro Ile Glu Ile A1a Asp Glu Thr
365 370 375
Thr Trp Ile Lys Glu Pro Val Asp Asn Gly Ser Leu Pro Met Thr
380 385 390
Asp Phe Val Ser Gly I1e Ala Phe Ser Ile Glu Asn Ser His Met
395 400 405
Cys Met Ser Pro Leu Ala Glu Ser Ser Val I1e Pro Cys Glu Ser
410 415 420
Ser Asn Ile Gln Met Asp Ser Gly Tyr Asn Thr Gln Asn Cys Gly
425 430 435
Ser Asn Ile Met Asp Thr Val Gly Ala Glu Ser Tyr Cys Lys Glu
440 445 450
Ser Asp Ala Gln Thr Cys Glu Val Glu Ser Lys Ser Gln A1a Phe
455 460 465
Asn Met Lys Gln Asp His Thr Thr Gln Arg Cys Trp Met Lys Thr
470 475 480
Ala Ser Pro Phe Gln Cys Ser Ser Pro
485
<210> 30
<211> 184
<212> PRT
<213> Homo sapiens
<220>
<221> misc_f eature
<223> Incyte ID No: 2369350CD1
<400> 30
Met Ser Asn Glu Arg Gly Phe Glu Asn Val Glu Leu Gly Va1 I1e
1 5 10 15
Gly Lys Lys Lys Lys Val Pro Arg Arg Val Ile His Phe Val Ser
20 25 30
Gly Glu Thr Met Glu G1u Tyr Ser Thr Asp G1u Asp Glu Val Asp
35 40 45
Gly Leu Glu Lys Lys Asp Val Leu Pro Thr Val Asp Pro Thr Lys
50 55 60
Leu Thr Trp Gly Pro Tyr Leu Trp Phe Tyr Met Leu Arg Ala Ala
65 70 75
Thr Ser Thr Leu Ser Va1 Tyr Asp Phe Leu Gly Glu Lys Ile Ala
80 85 90
Ser Val Leu G1y Tle Ser Thr Pro Lys Tyr Gln Tyr Ala Ile Asp
95 100 105
Glu Tyr Tyr Arg Met Lys Lys Glu Glu Glu Glu Glu Glu Glu Glu
110 115 120
Asn Arg Met Ser Glu Glu Ala Glu Lys G1n Tyr Gln Gln Asn Lys
125 130 135
Leu Gln Thr Asp Ser Ile Val Gln Thr Asp Gln Pro Glu Thr Val
140 145 150
Ile Ser Ser Ser Phe Val Asn Val Asn Phe Glu Met G1u Gly Asp
155 160 165
Ser Glu Val Ile Met Glu Ser Lys Gln Ile Gln Ser Leu Ser His
170 175 180
His Lys Met Lys
<210> 31
<211> 52,0
34/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<212> PRT
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 2722979CD1
<400> 31
Met Leu Gln Ile Thr Glu Trp Arg Phe Leu Ala Arg Asp Glu Gly
1 5 10 15
Glu Ser A1a Val A1a Glu Asp Pro Thr Trp Gly Glu Asp Glu Glu
20 25 30
Pro Ser Ala Cys Thr Thr Asp Ser Trp Ala G1n Gly Ser Val Pro
35 40 45
Val Leu His Ala Ser Thr Ser Glu Gly Leu Glu Asn Phe Gln Gly
50 55 60
Glu Va1 His Ser Ser Gly Ala Ser Pro Asp Ser Ser Ala Ile Ala
65 70 75
Pro Ala Leu Pro Phe Pro Thr Ser His Cys Pro Ser Ala Phe Pro
80 85 90
Gln Asp Pro Gly Gly Val Asp Arg Ile Pro Leu Gly Arg Ser Trp
95 100 105
Met Gly Arg Gly Ser Gln G1u Gln Met Glu Ser Trp Glu Pro Ser
110 115 120
Pro Gln Leu Arg Val Thr Ser Ala Pro Pro Pro Thr Ser Glu Leu
125 130 135
Phe Gln Glu Ala Gly Pro Gly Gly Pro Val Glu Glu Ala Asp Gly
140 145 150
Gln Ser Arg G1y Leu Ser Ser Ala Gly Ser Leu Ser Ala Ser Phe
155 160 165
Gln Leu Ser Val Glu Glu Ala Pro Ala Asp Asp Ala Asp Pro Ser
170 175 180
Leu Asp Pro Tyr Leu Val Ala Ser Pro Gln Ala Ser Thr Gly Arg
185 190 195
Gly His Pro Leu Gly Phe His Leu Ser Leu Glu Asp Leu Tyr Cys
200 205 210
Cys Met Pro Gln Leu Asp A1a Ala Gly Asp Arg Leu Glu Leu Arg
215 220 225
Ser Glu Gly Val Pro Cys Ile Ala Ser G1y Val Leu Val Ser Tyr
230 235 240
Pro Ser Val Gly Gly Ala Thr Arg Pro Ser Ala Ser Cys Gln G1n
245 250 255
Gln Arg Ala G1y His Ser Asp Va1 Arg Leu Ser Ala His His His
260 265 270
Arg Met Arg Arg Lys Ala Ala Val Lys Arg Leu Asp Pro Ala Arg
275 280 285
Leu Pro Cys His Trp Val Arg Pro Leu Ala Glu Val Leu Val Pro
290 295 300
Asp Ser Gln Thr Arg Pro Leu Glu Ala Tyr Arg Gly Arg Gln Arg
305 310 315
Gly Glu Lys Thr Lys A1a Arg Ala Glu Pro Gln Ala Leu Gly Pro
320 325 330
Gly Thr Arg Val Ser Pro Ala Ala Phe Phe Pro Leu Arg Pro Gly
335 340 345
Ile Pro Phe Arg Asp Leu Asp Ser Gly Pro Ala Leu Leu Phe Pro
350 355 360
Thr Leu Asn Leu Gly Leu Ser Ser Pro Ser Leu Glu Ser Lys Leu
365 370 375
Pro Leu Pro Asn Ser Arg Ile Arg Phe Leu Thr Thr His Pro Val
380 385 390
Leu Pro Asp Val Ala Arg Ser Arg Ser Pro Lys Leu Trp Pro Ser
395 400 405
Val Arg Trp Pro Ser Gly Trp Glu Gly Lys Ala G1u Leu Leu Gly
35/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
410 415 420
Glu Leu Trp Ala Gly Arg Thr Arg Va1 Pro Pro Gln G1y Leu Glu
425 430 435
Leu Ala Asp Arg Glu Gly Gln Asp Pro Gly Arg Trp Pro Arg Thr
440 445 450
Thr Pro Pro Val Leu Glu Ala Thr Ser Gln Val Met Trp Lys Pro
455 460 465
Val Leu Leu Pro Glu Ala Leu Lys Leu Ala Pro Gly Val Ser Met
470 475 480
Trp Asn Arg Ser Thr Gln Va1 Leu Leu Ser Ser Gly Va1 Pro Glu
485 490 495
Gln Glu Asp Lys Glu Gly Ser Thr Phe Pro Pro Val Glu Gln His
500 505 510
Pro Ile Gln Thr Gly Ala Pro Lys Pro Arg
515 520
<210> 32
<211> 255
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 60140470CD1
<400> 32
Met Ala Ser Ser Asp Leu Glu Gln Leu Cys Ser His Val Asn G1u
1 5 10 15
Lys Ile Gly Asn Ile Lys Lys Thr Leu Ser Leu Arg Asn Cys Gly
20 25 30
Gln Glu Pro Thr Leu Lys Thr Val Leu Asn Lys I1e Gly Asp Glu
35 40 ~ 45
Ile Ile Val Ile Asn Glu Leu Leu Asn Lys Leu Glu Leu Glu Ile
50 55 60
Gln Tyr Gln Glu Gln Thr Asn Asn Ser Leu Lys Glu Leu Cys Glu
65 70 75
Ser Leu Glu Glu Asp Tyr Lys Asp Ile Glu His Leu Lys Glu Asn
80 85 . 90
Val Pro Ser His Leu Pro Gln Val Thr Val Thr Gln Ser Cys Val
95 100 105
Lys Gly Ser Asp Leu Asp Pro Glu Glu Pro Ile Lys Val Glu Glu
110 115 120
Pro Glu Pro Val Lys Lys Pro Pro Lys Glu Gln Arg Ser Ile Lys
125 130 135
Glu Met Pro Phe I1e Thr Cys Asp Glu Phe Asn Gly Val Pro Ser
140 145 150
Tyr Met Lys Ser Arg Leu Thr Tyr Asn Gln Ile Asn Asp Val Ile
155 160 165
Lys Glu Ile Asn Lys Ala Val Ile Ser Lys Tyr Lys Ile Leu His
17f 175 180
Gln Pro Lys Lys Ser Met Asn Ser Val Thr Arg Asn Leu Tyr His
185 190 195
Arg Phe Ile Asp Glu Glu Thr Lys Asp Thr Lys Gly Arg Tyr Phe
200 205 210
Ile Val Glu Ala Asp Ile Lys Glu Phe Thr Thr Leu Lys Ala Asp
215 220 225
Lys Lys Phe His Val Leu Leu Asn Ile Leu Arg His Cys Arg Arg
230 ~ 235 240
Leu Ser Glu Val Arg Gly Gly Gly Leu Thr Arg Tyr Val 21e Thr
245 250 255
<210> 33
36/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<211> 231
<212> PRT
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 70623603CD1
<400> 33
Met Glu Asp Ser Pro Leu Pro Asp Leu Arg Asp Ile Glu Leu Lys
1 5 10 15
Leu Gly Arg Lys Val Pro Glu Ser Leu Val Arg Ser Leu Arg Gly
20 25 30
Glu Glu Pro Val Pro Arg Glu Arg Asp Arg Asp Pro Cys Gly Gly
35 40 45
Ser Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Cys Ser
50 55 60
Ser Gly Ser Ser Tyr Cys Ser Phe Pro Pro Ser Leu Ser Ser Ser
65 70 75
Ser Ser Ser Ser Pro Thr Ser Gly Ser Pro Arg G1y Ser His Ser
80 85 90
Ser A1a Leu Glu Arg Leu Glu Thr Lys Leu His Leu Leu Arg Gln
95 100 105
Glu Met Val Asn Leu Arg Ala Thr Asp Val Arg Leu Met Arg Gln
110 115 120
Leu Leu Val Ile Asn Glu Ser Ile Glu Ser Ile Lys Trp Met Ile
125 130 135
Glu Glu Lys Ala Thr I1e Thr Ser Arg G1y Ser Ser Leu Ser Gly
140 145 150
Ser Leu Cys Ser Leu Leu Glu Ser Gln Ser Thr Ser Leu Arg Gly
155 160 165
Ser Tyr Asn Ser Leu His Asp Gly Ser Asp G1y Leu Asp Gly Ile
170 175 180
Ser Va1 Gly Ser Tyr Leu Asp Thr Leu Ala Asp Asp Val Pro Gly
185 190 195
His Gln Thr Pro Ser Asp Leu Asp Gln Phe Ser Asp Ser Ser Leu
200 205 210
I1e Glu Asp Ser Gln Ala Leu His Lys Arg Pro Lys Leu Asp Ser
215 220 225
Glu Tyr Tyr Cys Phe Gly
230
<210> 34
<211> 492
<212> PRT
<213> Homo sapiens
<220>
<221> misC_f eature
<223> InCyte ID No: 7161479CD1
<400> 34
Met Leu Pro Ala Arg Trp Phe Cys Tyr Tyr Asn Ser His Pro Lys
1 5 10 15
Tyr Arg Arg Cys Ser Val Pro Glu Glu Gln Glu Leu Thr Asp Glu
~20 25 30
Asp Leu Cys Leu Ser Lys Ala Lys Lys Gln Glu Gln Thr Val Glu
35 40 45
Glu Lys Lys Lys Met Pro Met Glu Asn Glu Asn His Gln Val Phe
50 55 60
Ser Asn Pro Pro Lys Ile Leu Thr Val G1n Glu Met Ala G1y Leu
65 70 75
Asn Asn Lys Thr Ile Gly Tyr Glu Gly Ile His Ser Pro Ser Val
37/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
80 85 90
Leu Pro Ser G1y Gly Glu Glu Ser Arg Ser Pro Ser Leu Gln Leu
95 100 105
Lys Pro Leu Asp Ser Ser Val Leu G1n Phe Ser Ser Lys Tyr Lys
110 115 120
Trp I1e Leu Gly Glu Glu Pro Val Glu Lys Arg Arg Arg Leu Gln
125 ~ 130 135
Asn G1u Met Thr Thr Pro 'Ser Leu Asp Tyr Ser Met Pro Ala Pro
140 145 150
Tyr Arg Arg Val Glu Ala Pro Val Ala Tyr Pro Glu Gly Glu Asn
155 160 165
Ser His Asp Lys Ser Ser Ser Glu Arg Ser Thr Pro Pro Tyr Leu
170 175 180
Phe Pro Glu Tyr Pro Glu Ala Ser Lys Asn Thr Gly Gln Asn Arg
185 190 195
Glu Va1 Ser Ile Leu Tyr Pro Gly Ala Lys Asp G1n Arg G1n Gly
200 205 210
Ser Leu Leu Pro Glu Glu Leu Glu Asp Gln Met Pro Arg Leu Val
215 220 225
Ala Glu Glu Ser Asn Arg G1y Ser Thr Thr Ile Asn Lys Glu Glu
230 . 235 240
Val Asn Lys Gly Pro Phe Val Ala Val Val Gly Val Ala Lys Gly
245 250 255
Val Arg Asp Ser Gly Ala Pro Ile Gln Leu Ile Pro Phe Asn Arg
260 265 270
Glu Glu Leu Ala G1u Arg Arg Lys A1a Val Glu Ser Trp Asn Pro
275 280 285
Val Pro Tyr Ser Val Ala Ser A1a Ala Ile Pro Ala Ala Ala Ile
290 295 300
Gly Glu Lys Ala Arg Gly Tyr Glu Glu Ser Glu Gly His Asn Thr
305 310 315
Pro Lys Leu Lys Asn Gln Arg Glu Leu Glu Glu Leu Lys Arg Thr
320 325 330
Thr G1u Lys Leu Glu Arg Val Leu Ala Glu Arg Asn Leu Phe Gln
335 340 345
Gln Lys Val Glu Glu Leu Glu Gln Glu Arg Asn His Trp Gln Sex
350 355 360
Glu Phe Lys Lys Val Gln His G1u Leu Val Ile Tyr Ser Thr Gln
365 370 375
Glu Ala Glu Gly Leu Tyr Trp Ser Lys Lys His Met Gly Tyr Arg
380 385 390
Gln Ala Glu Phe Gln Ile Leu Lys Ala Glu Leu Glu Arg Thr Lys
395 400 405
Glu Glu Lys G1n Glu Leu Lys Glu Lys Leu Lys Glu Thr Glu Thr
410 415 420
His Leu Glu Met Leu Gln Lys Ala Gln Val Ser Tyr Arg Thr Pro
425 430 435
Glu Gly Asp Asp Leu Glu Arg Ala Leu Ala Lys Leu Thr Arg Leu
440 445 450
Arg Ile His Va1 Ser Tyr Leu Leu Thr Ser Va1 Leu Pro His Leu
455 460 465
Glu Leu Arg G1u Ile Gly Tyr Asp Ser Glu Gln Val Asp Gly Ile
470 475 480
Leu Tyr Thr Val Leu Glu Ala Asn His Ile Leu Asp
485 490
<210> 35
<211> 85
<212> PRT
<213> Homo sapiens
<220>
<221> misc_feature
38/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<223> Inoyte ID No: 7502313CD1
<400> 35
Met Ser Leu Phe His Arg Asn Val Asn Leu Thr Val Thr Ser Glu
1 5 10 15
Phe Val Gln Cys Pro Thr Met Val Tyr Glu Lys Tyr Thr Gly Ser
20 25 30
Val Gly Gly Thr His Asp Met Ile Cys Glu Tyr His His Leu Cys
35 40 45
Gln Thr Ser Leu Gln Gly Ile Pro Val Ser Gln Leu Lys Gly Val
50 55 60
Asn Gly His Thr His Ser Leu Asp Asp A1a Leu Ala Val Leu Arg
65 70 75
Gly Cys Lys Val Gly Ser Gly Pro Ser Ser
80 85
<210> 3 6
<211> 178
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7502390CD1
<400> 36
Met Gly Glu Thr Trp Lys Asn Ile Cys Ser Thr Val Arg His Gly
1 5 10 15
Trp Trp Leu Arg Asp His Arg Met A1a Gly Leu Pro Ile Pro Pro
20 25 30
Glu Tle Val Lys Glu Ala Glu Val Pro Gln A1a Ala Leu Gly Val
35 40 45
Pro Ala Gln Gly Thr Gly Asp Asn Gly His Thr Pro Val Glu Glu
50 55 60
Glu Va1 Gly Gly Ile Pro Val Pro Ala Pro Gly Leu Leu Gln Val
65 70 75
Thr Glu Arg Arg Gln Pro Leu Ser Ser Val Ser Ser Leu Glu Va1
80 85 90
His Phe Asp Leu Leu Asp Leu Thr Glu Leu Thr Asp Met Ser Asp
95 100 105
Gln Glu Leu Ala Glu Va1 Phe Ala Asp Ser Asp Asp Glu Asn Leu
110 115 120
Asn Thr Glu Ser Pro A1a Gly Leu His Pro Leu Pro Arg Ala Gly
125 130 135
Tyr Leu Arg Ser Pro Ser Trp Thr Arg Thr Arg Ala Glu Gln Ser
140 145 150
His Glu Lys Gln Pro Leu Gly Asp Pro Glu Arg Gln Ala Thr Val
155 160 165
Leu Asp Thr Phe Leu Thr Val Glu Arg Pro Gln G1u Asp
170 175
<210> 37
<211> 665
<212> PRT
<213> Homo sapiens
<220>
<221> misC_feature
<223> Incyte ID No: 7502872CD1
<400> 37
Met Gly Asp Ile Leu Ala His Glu Ser Glu Leu Leu Gly Leu Val
1 5 10 15
39/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
Lys G1u Tyr Leu Asp Phe Ala Glu Phe Glu Asp Thr Leu Lys Thr
20 25 30
Phe Ser Lys Glu Cys Lys Ile Lys Gly Lys Pro Leu Cys Lys Thr
35 40 45
Val Gly Gly Ser Phe Arg Asp Ser Lys Ser Leu Thr Ile Gln Lys
50 55 60
Asp Leu Val Ala Ala Phe Asp Asn Gly Asp Gln Lys Val Phe Phe
65 70 75
Asp Leu Trp Glu Glu His Ile Ser Ser Ser Ile Arg Asp Gly Asp
80 85 90
Ser Phe Ala Gln Lys Leu Glu Phe Tyr Leu His Ile His Phe Ala
95 100 105
Ile Tyr Leu Leu Lys Tyr Ser Val Gly Arg Pro Asp Lys Glu Glu
110 115 120
Leu Asp Glu Lys Ile Ser Tyr Phe Lys Thr Tyr Leu Glu Thr Lys
125 130 135
Gly Ala Ala Leu Ser Gln Thr Thr G1u Phe Leu Pro Phe Tyr Ala
140 145 ' 150
Leu Pro Phe Val Pro Asn Pro Met Val His Pro Ser Phe Lys Glu
155 160 165
Leu Phe Gln Asp Ser Trp Thr Pro Glu Leu Lys Leu Lys Leu Glu
170 175 180
Lys Phe Leu Ala Leu Ile Ser Lys Ala Ser Asn Thr Pro Lys Leu
185 190 195
Leu Thr Ile Tyr Lys Glu Asn Gly G1n Ser Asn Lys Glu Ile Leu
200 205 210
Gln Gln Leu His Gln G1n Leu Va1 G1u Ala Glu Arg Arg Ser Va1
215 220 225
Thr Tyr Leu Lys Arg Tyr Asn Lys Ile Gln Ala Asp Tyr His Asn
230 235 240
Leu Ile Gly Val Thr Ala Glu Leu Val Asp Ser Leu Glu Ala Thr
245 250 255
Va1 Ser Gly Lys Met Ile Thr Pro Glu Tyr Leu Gln Ser Val Cys
260 265 270
Val Arg Leu Phe Ser Asn Gln Met Arg Gln Ser Leu A1a His Ser
275 280 285
Val Asp Phe Thr Arg Pro Gly Thr Ala Ser Thr Met Leu Arg Ala
290 295 300
Ser Leu Ala Pro Val Lys Leu Lys Asp Val Pro Leu Leu Pro Ser
305 310 315
Leu Asp Tyr Glu Lys Leu Lys Lys Asp Leu Ile Leu Gly Ser Asp
320 325 330
Arg Leu Lys Ala Phe Leu Leu Gln Ala Leu Arg Trp Arg Leu Thr
335 340 345
Thr Ser His Pro Gly Glu Gln Arg Glu Thr Val Leu G1n Ala Tyr
350 355 360
Ile Ser Asn Asp Leu Leu Asp Cys Tyr Ser His Asn Gln Arg Ser
365 370 375
Val Leu Gln Leu Leu His Ser Thr Ser Asp Val Val Arg Gln Tyr
380 385 390
Met Ala Arg Leu Ile Asn Ala Phe Ala Ser Leu Ala Glu Gly Arg
395 . 400 405
Leu Tyr Leu Ala Gln Asn Thr Lys Val Leu Gln Met Leu Glu Gly
410 415 420
Arg Leu Lys Glu Glu Asp Lys Asp Ile Ile Thr Arg Glu Asn Va1
425 430 435
Leu Gly Ala Leu Gln Lys Phe Ser Leu Arg Arg Pro Leu Gln Thr
440 445 450
A1a Met I1e Gln Asp G1y Leu Ile Phe Trp Leu Val Asp Val Leu
455 460 465
Lys Asp Pro Asp Cys Leu Ser Asp Tyr Thr Leu Glu Tyr Ser Val
470 475 480
Ala Leu Leu Met Asn Leu Cys Leu Arg Ser Thr Gly Lys Asn Met
40/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
485 490 495
Cys A1a Lys Val Ala Gly Leu Val Leu Lys Val Leu Ser Asp Leu
500 505 510
Leu Gly His Glu Asn His Glu I1e Gln Pro Tyr Va1 Asn Gly A1a
515 520 525
Leu Tyr Ser Ile Leu Ser Val Pro Ser I1e Arg Glu Glu Ala Arg
530 535 540
Ala Met Gly Met Glu Asp Ile Leu Arg Cys Phe Ile Lys Glu Gly
545 550 555
Asn Ala Glu Met Ile Arg Gln Ile Glu Phe I1e Ile Lys Gln Leu
560 565 570
Asn Ser Glu Glu Leu Pro Asp Gly Val Leu Glu Ser Asp Asp Asp
575 580 585
Glu Asp Glu Asp Asp Glu Glu Asp His Asp Ile Met Glu Ala Asp
590 595 600
Leu Asp Lys Asp Glu Leu Ile Gln Pro Gln Leu Gly Glu Leu Ser
605 610 615
Gly Glu Lys Leu Leu Thr Thr Glu Tyr Leu Gly Ile Met Thr Asn
620 625 630
Thr Gly Lys Thr Arg Arg Lys Gly Leu A1a Asn Val Gln Trp Ser
635 640 645
Gly Asp Glu Pro Leu Gln Arg Pro Val Thr Pro Gly Gly His Arg
650 655 660
Asn Gly Tyr Pro Val
665
<210> 38
<211> 551
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7505443CD1
<400> 38
Met Ala Arg Ala Gly Pro Arg Leu Val Leu Ser G1u G1u A1a Val
1 5 10 15
Arg Ala Lys Ser Gly Leu G1y Pro His Arg Asp Leu A1a Glu Leu
20 25 30
Gln Ser Leu Ser Ile Pro Gly Thr Tyr Gln Glu Lys Ile Thr His
35 40 45
Leu Gly His Ser Leu Met Ser Leu Thr G1y Leu Lys Ser Leu Asp
50 55 60
Leu Ser Arg Asn Ser Leu Val Ser Leu Glu Gly Ile Gln Tyr Leu
65 70 75
Thr Ala Leu Glu Ser Leu Asn Leu Tyr Tyr Asn Cys Ile Ser Ser
80 85 90
Leu Ala Glu Val Phe Arg Leu His Ala Leu Thr Glu Leu Val Asp
95 100 105
Val Asp Phe Arg Leu Asn Pro Val Val Lys Val Glu Pro Asp Tyr
110 115 120
Arg Leu Phe Val Val His Leu Leu Pro Lys Leu Gln Gln Leu G1u
125 130 135
Ser Arg His Leu Leu Ser Pro Gln Leu Val Gln Tyr Gln Cys G1y
140 145 150
Asp Ser Gly Lys G1n Gly Arg Glu Thr Arg Arg Ser Ser Cys Arg
155 160 165
G1y Cys Cys Leu Glu Lys Met Pro Trp Ser Gln Leu Cys Gly Glu
170 175 180
Leu Pro Pro Leu Tyr Gly Ala Glu Pro Glu Ala Ser Arg Ala Pro
185 190 195
Arg Pro His Thr Tyr Phe Thr Pro His Pro Asp Ser Met Asp Thr
41/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
200 205 210
Glu Asp Ser Ala Ser Ser Gln Lys Leu Asp Leu Ser Gly Glu Met
215 220 225
Val Pro Gly Pro Leu Pro Ala Pro G1y Lys Cys Arg Lys Arg Arg
230 ~ 235 240
Met Pro Va1 Gly Arg Phe Gln Thr Phe Ser Asp G1n Glu Gly Leu
245 250 255
Gly Cys Pro Glu Arg Thr His Gly Ser Ser Val Pro Lys Glu Ser
260 265 270
Leu Ser Arg Gln Asp Ser Ser Glu Ser Arg Asn Gly Arg Thr Leu
275 280 285
Ser Gln Pro Glu Ala Ser Glu Thr Glu Glu Gln Arg Ser Arg Gly
290 295 300
Va1 Thr Asp Thr Arg G1u Pro Ser Pro Gly Ser His Ser Ala Leu
305 310 315
Pro Gly Lys Lys Thr Ala Leu Gln Ala Ala Leu Leu Glu Thr Leu
320 325 330
Leu Asp Leu Val Asp Arg Ser Trp Gly Gly Cys Arg Ser Leu His
335 340 345
Ser Asn Glu Ala Phe Leu A1a Gln A1a Arg His Ile Leu Ser Ser
350 355 360
Val Glu Glu Phe Thr A1a Ala Gln Asp Ser Ser Ala Met Val Gly
365 370 375
Glu Asp Val Gly Ser Leu A1a Leu Glu Ser Lys Ser Leu Gln Ser
380 385 390
Arg Leu Ala Glu Gln G1n Gln Gln His Ala Arg Glu Met Ser Glu
395 400 405
Val Thr Ala Glu Leu His His Ala His Lys G1u Leu Asp Asp Leu
410 415 420
Arg Gln His Leu Asp Lys Ser Leu Glu Glu Asn Ser Arg Leu Lys
425 430 435
Ser Leu Trp Leu Ser Met Lys Lys G1u Val Lys Ser Ala Asp Thr
440 445 450
Ala Ala Thr Leu Asn Leu .Gln Ile Ala Gly Leu Gln Thr Ser Val
455 460 465
Lys Arg Leu Cys Gly G1u Ile Val Glu Leu Lys Gln His Leu Glu
470 475 480
His Tyr Asp Lys Ile Gln Glu Leu Thr Gln Met Leu Gln Glu Ser
485 490 495
His Ser Ser Leu Val Ser Thr Asn G1u His Leu Leu Gln Glu Leu
500 505 510
Ser G1n Val Arg Ala G1n His Arg Ala Glu Val Glu Gln Met His
515 520 525
Trp Ser Tyr Gln Glu Leu Lys Lys Thr Met Ala Leu Phe Pro His
530 535 540
Ser Ser Ala Ser His Gly Gly Cys Gln Ala Cys
545 550
<210> 39
<211> 148
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID I~7o: 8032443CD1
<400> ~39
Met Gly Pro Glu Glu Lys Thr Ile Met Thr Glu Arg Ser Ala Ala
1 5 10 15
Va1 Phe Ile Gln Ala Trp Trp Arg Gly Met Leu Va1 Arg Arg Thr
20 25 30
Leu Leu His Ala Ala Leu Arg Ala Trp Ile Ile Gln Cys Trp Trp
42190
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
35 40 45
Arg Gln Va1 Leu Glu Lys Leu Leu Ala Lys Arg Arg Arg Met Val
50 55 60
Leu G1u Phe Tyr Val Gln G1n Glu Trp Ala Ala Val Arg Leu Gln
65 70 75
Ser Trp Va1 Arg Met Trp Cys Val Arg Gln Arg Tyr Cys Arg Leu
80 85 90
Leu Asn Ala Val Arg Ile Ile Gln Val Tyr Trp Arg Trp His Ser
95 100 105
Cys His Ser Arg Val Phe Ile Glu G1y His Tyr Glu Leu Lys Glu
110 115 120
Asn Gln Leu Asn Ile Gln Leu G1u Ile Ser Leu Gly Leu Gln Ala
125 130 135
Cys Lys Val G1n Gln Cys Ile Pro Leu Pro Leu Lys Glu
140 145
<210> 40
<211> 342
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7704916CD1
<400> 40
Met Lys Thr Val Lys Glu Lys Lys Glu Cys Gln Arg Leu Arg Lys
1 5 10 15
Ser A1a Lys Thr Arg Arg Val Thr G1n Arg Lys Pro Ser Ser G1y
20 25 30
Pro Val Cys Trp Leu Cys Leu Arg Glu Pro Gly Asp Pro Glu Lys
35 40 45
Leu Gly Glu Phe Leu Gln Lys Asp Asn Ile Ser Val His Tyr Phe
50 55 60
Cys Leu Ile Leu Ser Ser Lys Leu Pro Gln Arg Gly Gln Ser Asn
65 70 75
Arg Gly Phe His G1y Phe Leu Pro Glu Asp Ile Lys Lys Glu Ala
80 85 90
Ala Arg Ala Ser Arg Lys Ile Cys Phe Val Cys Lys Lys Lys Gly
95 100 105
Ala Ala Ile Asn Cys Gln Lys Asp Gln Cys Leu Arg Asn Phe His
110 115 120
Leu Pro Cys Gly Gln Glu Arg Gly Cys Leu Ser Gln Phe Phe G1y
125 130 135
Glu Tyr Lys Ser Phe Cys Asp Lys His Arg Pro Thr Gln Asn Ile
140 145 150
Gln His Gly His Val Gly Glu Glu Ser Cys Ile Leu Cys Cys Glu
155 160 165
Asp Leu Ser Gln Gln Ser Val Glu Asn Ile Gln Ser Pro Cys Cys
170 175 180
Ser Gln Ala Ile Tyr His Arg Lys Cys I1e Gln Lys Tyr Ala His
185 190 195
Thr Ser Ala Lys His Phe Phe Lys Cys Pro Gln Cys Asn Asn Arg
200 205 210
Lys Glu Phe Pro Gln Glu Met Leu Arg Met Gly Ile His Ile Pro
215 220 225
Asp Arg Arg Trp Cys Leu Ile Leu Cys Ala Thr Cys Gly Ser His
230 235 240
Gly Thr His Arg Asp Cys Ser Ser Leu Arg Ser Asn Ser Lys Lys
245 250 255
Trp Glu Cys G1u Glu Cys Ser Pro A1a A1a Ala Thr Asp Tyr Ile
260 265 270
Pro G1u Asn Ser Gly Asp Ile Pro Cys Cys Ser Ser Thr Phe His
43/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
275 280 285
°Pro Glu Glu His Phe Cys Arg Asp Asn Thr Leu Glu G1u Asn Pro
290 295 300
Gly Leu Ser Trp Thr Asp Trp Pro Glu Pro Ser Leu Leu Glu Lys
305 310 315
Pro Glu Ser Ser Arg Gly Arg Arg Ser Tyr Ser Trp Arg Ser Lys
320 325 330
G1y Val Arg Ile Thr Asn Ser Cys Lys Lys Ser Lys
335 340
<210> 41
<211> 194
<212> PRT
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 2013440CD1
<400> 41
Met Phe Leu Thr Ala Val Asn Pro Gln Pro Leu Ser Thr Pro Ser
1 5 10 15
Trp Gln Ile Glu Thr Lys Tyr Ser Thr Lys Val Leu Thr Gly Asn
20 25 30
Trp Met Glu Glu Arg Arg Lys Phe Thr Arg Asp Thr Asp Lys Thr
35 40 45
Pro Gln Ser Ile Tyr Arg Lys Glu Tyr Ile Pro Phe .Pro Asp His
50 55 60
Arg Pro Asp Gln Ile Ser Arg Trp Tyr Gly Lys Arg Lys Val Glu
65 70 75
Gly Leu Pro Tyr Lys His Leu Ile Thr His His Gln Glu Pro Pro
80 85 90
His Arg Tyr Leu Ile Ser Thr Tyr Asp Asp His Tyr Asn Arg His
95 100 105
Gly Tyr Asn Pro Gly Leu Pro Pro Leu Arg Thr Trp Asn G1y Gln
110 115 120
Lys Leu Leu Trp Leu Pro Glu Lys Ser Asp Phe Pro Leu Leu Ala
125 130 135
Pro Pro Thr Asn Tyr Gly Leu Tyr Glu Gln Leu Lys Gln Arg Gln
140 145 150
Leu Thr Pro Lys Ala Gly Leu Lys Gln Ser Thr Tyr Thr Ser Ser
155 160 165
Tyr Pro Arg Pro Pro Leu Cys Ala Met Ser Trp Arg Glu His Ala
170 175 180
Val Pro Val Pro Pro His Arg Leu His Pro Leu Pro His Phe
185 190
<210> 42
<211> 126
<212> PRT
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 2503512CD1
<400> 42
Met Gly Gly Gln Ala Gly Leu Arg Thr Gly Arg Thr Lys Arg Gly
1 5 10 15
Glu Glu Arg Pro His Thr Cys Ser Asp Ile Lys Ser Ile Leu Leu
20 25 30
His Arg Tyr Phe Arg Cys Gln Gly Leu Gln Ala Gly Ser Pro G1n
35 40 45
44/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
Leu Leu Pro G1y Thr Ser Pro Thr Gly Asp Cys Arg Ala Leu Gly
50 55 60
Trp Val Thr Pro Pro Ala Pro Arg Lys Thr Ser Ser Leu Ala Thr
65 70 75
Pro Arg Pro Leu Ser Ser Lys Gln Ser Ala Arg Ser Ser Ser Gly
80 85 90
Ser Pro Arg Asn Arg Ala Pro Cys Arg Thr Ser Thr Ala Asp Arg
95 100 105
Pro Arg Leu Ala Asp Leu Pro Ser Ile Arg Phe Leu Trp Lys Gln
110 115 120
Asp Gln Lys Glu Ile Asn
125
<210> 43
<211> 474
<212> PRT
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 277396CD1
<400> 43
Met Lys Thr Lys Cys Ile Cys Glu Leu Cys Ser Cys Gly Arg His
1 ~ 5 10 15
His Cys Pro His Leu Pro Thr Arg Ile Tyr Asp Glu Thr Glu Lys
20 25 30
Pro Cys Leu Leu Ser G1u Tyr Thr Glu Asn Tyr Pro Phe Tyr His
35 40 45
Ser Tyr Leu Pro Arg Glu Ser Phe Lys Pro Arg Arg Glu Tyr G1n
50 55 60
Lys Gly Ser Ile Pro Met Glu G1y Leu Thr Thr Ser Arg Arg Asp
65 70 75
Phe Gly Pro His Lys Val A1a Pro Val Lys Va1 His Gln Tyr Asp
80 85 90
Gln Phe Va1 Pro Ser Glu Glu Asn Met Asp Leu Leu Thr Thr Tyr
95 100 105
Lys Lys Asp Tyr Asn Pro Tyr Pro Val Cys Arg Va1 Asp Pro Ile
110 115 120
Lys Pro Arg Asp Ser Lys Tyr Pro Cys Ser Asp Lys Met Glu Cys
125 130 135
Leu Pro Thr Tyr Lys Ala Asp Tyr Leu Pro Trp Asn Gln Pro Arg
140 145 150
Arg Glu Pro Leu Arg Leu G1u His Lys Tyr Gln Pro Ala Ser Va1
155 160 165
Arg Phe Asp Asn Arg Thr Thr His Gln Asp Asp Tyr Pro Ile Lys
170 175 180
Gly Leu Val Lys Thr Ile Ser Cys Lys Pro Leu Ala Met Pro Lys
185 190 195
Leu Cys Asn Ile Pro Leu Glu Asp Val Thr Asn Tyr Lys Met Ser
200 205 210
Tyr Va1 Ala His Pro Val G1u Lys Arg Phe Val His Glu Ala Glu
215 220 225
Lys Phe Arg Pro Cys Glu Ile Pro Phe Glu Ser Leu Thr Thr Gln
230 235 240
Lys Gln Ser Tyr Arg Gly Leu Met Gly Glu Pro Ala Lys Ser Leu
245 250 255
Lys Pro Leu Ala Arg Pro Pro G1y Leu Asp Met Pro Phe Cys Asn
260 265 270
Thr Thr Glu Phe Arg Asp Lys Tyr Gln Ala Trp Pro Met Pro Arg
275 280 285
Met Phe Ser Lys Ala Pro I1e Thr Tyr Val Pro Pro Glu Asp Arg
290 295 300
45/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
Met Asp Leu Leu Thr Thr Val Gln Ala His Tyr Thr Cys Pro Lys
305 310 315
Gly Ala Pro Ala Gln Ser Cys Arg Pro Ala Leu Gln Ile Lys Lys
320 325 330
Cys Gly Arg Phe Glu Gly Ser Ser Thr Thr Lys Asp Asp Tyr Lys
335 340 345
Gln Trp Ser Ser Met Arg Thr Glu Pro Val Lys Pro Val Pro Gln
350 355 360
Leu Asp Leu Pro Thr Glu Pro Leu Asp Cys Leu Thr Thr Thr Arg
365 370 375
Ala His Tyr Val Pro His Leu Pro Ile Asn Thr Lys Ser Cys Lys
380 385 390
Pro His Trp Ser Gly Pro Arg Gly Asn Val Pro Val Glu Ser Gln
395 400 405
Thr Thr Tyr Thr I1e Ser Phe Thr Pro Lys Glu Met G1y Arg Cys
410 415 420
Leu Ala Ser Tyr Pro G1u Pro Pro Gly Tyr Thr Phe Glu Glu Val
425 430 435
Asp Ala Leu Gly His Arg I1e Tyr Lys Pro Val Ser Gln Ala Gly
440 445 450
Ser G1n Gln Ser Ser His Leu Ser Val Asp Asp Ser Glu Asn Pro
455 460 465
Asn Gln Arg Glu Leu Glu Val Leu Ala
470
<210> 44
<211> 341
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 3044046CD1
<400> 44
Met Ser Val Leu Asp Ala Leu Trp Glu Asp Arg Asp Val Arg Phe
1 5 10 15
Asp Leu Ser A1a Gln Gln Met Lys Thr Arg Pro Gly Glu Val Leu
20 25 30
Ile Asp Cys Leu Asp Ser Ile Glu Asp Thr Lys Gly Asn Asn G1y
35 40 45
Asp Arg Gly Arg Leu Leu Val Thr Asn Leu Arg Ile Leu Trp His
50 55 60
Ser Leu Ala Leu Ser Arg Val Asn Val Ser Val Gly Tyr Asn Cys
65 70 75
Ile Leu Asn Ile Thr Thr Arg Thr Ala Asn Ser Lys Leu Arg Gly
80 85 90
Gln Thr Glu Ala Leu Tyr Ile Leu Thr Lys Cys Asn Ser Thr Arg
95 100 105
Phe Glu Phe Ile Phe Thr Asn Leu Val Pro Gly Ser Pro Arg Leu
110 115 120
Phe Thr Ser Val Met Ala Val His Arg Ala Tyr Glu Thr Ser Lys
125 130 135
Met Tyr Arg Asp Phe Lys Leu Arg Ser Ala Leu Ile Gln Asn Lys
140 145 150
Gln Leu Arg Leu Leu Pro Gln Glu His Val Tyr Asp Lys Ile Asn
155 160 165
Gly Val Trp Asn Leu Ser Ser Asp Gln Gly Asn Leu Gly Thr Phe
170 175 180
Phe Ile Thr Asn Val Arg Ile Val Trp His Ala Asn Met Asn Asp
185 190 195
Ser Phe Asn Val Ser Ile Pro Tyr Leu Gln Ile Arg Ser Ile Lys
200 205 210
46/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
I1e Arg Asp Ser Lys Phe Gly Leu Ala Leu Val Ile Glu Ser Ser
215 220 225
Gln Gln Ser Gly Gly Tyr Val Leu Gly Phe Lys Ile Asp Pro Val
230 235 240
Glu Lys Leu Gln Glu Ser Val Lys Glu Ile Asn Ser Leu His Lys
245 250 255
Val Tyr Ser Ala Ser Pro I1e Phe Gly Val Asp Tyr Glu Met Glu
260 265 270
Glu Lys Pro G1n Pro Leu Glu A1a Leu Thr Val Glu Gln Ile Gln
275 280 285
Asp Asp Val Glu Ile Asp Ser Asp Gly His Thr Asp Ala Phe Val
290 295 300
Ala Tyr Phe Ala Asp Gly Asn Lys Gln Gln Asp Arg Glu Pro Val
305 310 315
Phe Ser Glu Glu Leu Gly Leu Ala Ile G1u Lys Leu Lys Asp Gly
320 325 330
Phe Thr Leu Gln Gly Leu Trp Glu Val Met Ser
335 340
<210> 45
<211> 287
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 3808420CD1
<400> 45
I~et Asp Met His Ser Ala Arg Leu Asp Ser Phe Leu Ser Gln Leu
1 5 10 15
Arg Trp Glu Leu Leu Cys G1y Arg Asp Thr Gly Ser Pro Ser Met
20 25 30
Pro Gly Pro Leu Gln Pro Thr Ser Gln Thr Gly Pro Asp Val Gln
35 40 45
Pro Ser His Gln Leu Arg Ala Ser G1y Ala Leu Glu Glu Asp Ser
50 55 60
Val Cys Cys Val Glu G1u Glu G1u Glu Glu G1u Glu Glu Ala Val
65 70 75
Val Thr Glu Asp Arg Asp A1a Ala Leu Gly Gly Pro Arg Glu His
80 85 90
Ala Leu Asp Trp Asp Ser Gly Phe Ser Glu Va1 Ser Gly Ser Thr
95 100 105
Trp Arg G1u Glu Glu Leu Pro Val Ser Gln Arg Pro A1a Pro Ser
110 115 120
Ala Gln Pro Leu Arg Arg Gln Cys Leu Ser Val Ser Gly Leu Pro
125 130 135
Met Pro Ser Arg Ala Pro Val Ala Ser Val Pro Pro Val His His
140 145 150
Pro Arg Pro Lys Ser Thr Pro Asp Ala Cys Leu Glu His Trp Gln
155 160 165
Gly Leu Glu Ala Glu Asp Trp Thr Ala Ala Leu Leu Asn Arg Gly
170 175 180
Arg Ser Arg Gln Pro Leu Val Leu Gly Asp Asn Cys Phe Ala Asp
185 190 195
Leu Val His Asn Trp Met G1u Leu Pro Glu Thr Gly Ser Glu Gly
200 205 210
Gly Asp Gly Gly G1y His Arg Ala Arg Ala Arg Pro Pro Gln Phe
215 220 225
Leu Leu G1y Leu Ser Glu Gln Leu Arg Arg Arg Leu Ala Arg Ala
230 235 240
Arg Arg Thr Ala Met A1a Gly Lys Arg Leu Ser Cys Pro Pro Arg
245 250 255
47/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
Pro Glu Pro Glu Leu Pro Ala Asp Val Ser Arg Phe Ala Ala Leu
260 265 270
Met Ser Cys Arg Ser Arg Gln Pro Ile Ile Cys Asn Asp Val Ser
275 280 285
Tyr Leu
<210> 46
<211> 644
<212> PRT
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7504028CD1
<400> 46
Met Asn Leu Leu Pro Lys Ser Ser Arg Glu Phe Gly Ser Val Asp
1 5 10 15
Tyr Trp Glu Lys Phe Phe Gln Gln Arg Gly Lys Lys Ala Phe G1u
20 25 30
Trp Tyr Gly Thr Tyr Leu Glu Leu Cys Gly Val Leu His Lys Tyr
35 40 45
Ile Lys Pro Arg Glu Lys Val Leu Val Ile Gly Cys Gly Asn Ser
50 55 60
Glu Leu Ser G1u Gln Leu Tyr Asp Va1 Gly Tyr Arg Asp Ile Val
65 70 75
Asn Ile Asp Ile Ser Glu Val Va1 Ile Lys Gln Met Lys Glu Cys
80 85 90
Asn Ala Thr Arg Arg Pro Gln Met Ser Phe Leu Lys Met Asp Val
95 100 105
Thr Gln Met Glu Phe Pro Asp A1a Ser Phe G1n Val Val Leu Asp
110 115 120
Lys Gly Thr Leu Asp Ala Val Leu Thr Asp Glu Glu Glu Lys Thr
125 130 135
Leu Gln Gln Val Asp Arg Met Leu Ala Glu Val Gly Arg Val Leu
140 145 150
Gln Val Gly Gly Arg Tyr Leu Cys Ile Ser Leu Ala Gln A1a His
155 160 165
Ile Leu Lys Lys Ala Val Gly His Phe Ser Arg G1u G1y Trp Met
170 175 180
Va1 Arg Val His Gln Val Ala Asn Ser Gln Asp Gln Va1 Leu Glu
185 190 195
Ala Glu Pro Gln Phe Ser Leu Pro Val Phe Ala Phe Ile Met Thr
200 205 210
Lys Phe Arg Pro Va1 Pro G1y Ser Ala Leu Gln Ile Phe Glu Leu
215 220 225
Cys Ala Gln Glu Gln Arg Lys Pro Val Arg Leu G1u Ser A1a Glu
230 235 240
Arg Leu Ala Glu Ala Val Gln Glu Arg Gln Gln Tyr Ala Trp Leu
245 250 255
Cys Ser Gln Leu Arg Arg Lys Ala Arg Leu Gly Ser Val Ser Leu
260 265 270
Asp Leu Cys Asp Gly Asp Thr Gly G1u Pro Arg Tyr Thr Leu His
A 275 280 285
Val Val Asp Ser Pro Thr Val Lys Pro Ser Arg Asp Asn His Phe
290 295 300
Ala Ile Phe Ile Ile Pro Gln Gly Arg Glu Thr Glu Trp Leu Phe
305 310 315
Gly Met Asp Glu Gly Arg Lys Gln Leu Ala Ala Ser Ala Gly Phe
320 325 330
Arg Arg Leu Ile Thr Val Ala Leu His Arg Gly Gln Gln Tyr Glu
335 340 345
48/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
Ser Met Asp His I1e Gln Ala Glu Leu Ser Ala Arg Val Met Glu
350 355 360
Leu Ala Pro Ala Gly Met Pro Thr Gln Gln Gln Val Pro Phe Leu
365 370 375
Ser Val Gly Gly Asp Ile Gly Val Arg Thr Val Gln His Gln Asp
380 385 390
Cys Ser Pro Leu Ser Gly Asp Tyr Val I1e Glu Asp Val Gln Gly
395 400 405
Asp Asp Lys Arg Tyr Phe Arg Arg Leu I1e Phe Leu Ser Asn Arg
410 415 420
Asn Val Val Gln Ser Glu Ala Arg Leu Leu Lys Asp Val Ser His
425 430 435
Lys Glu Ile Pro Leu Ala Leu Leu Val Val Gly Leu Gly Gly Gly
440 445 450
Ser Leu Pro Leu Phe Val His Asp His Phe Pro Lys Ser Cys Ile
455 460 465
Asp Ala Val Glu Ile Asp Pro Ser Met Leu Glu Va1 Ala Thr Gln
470 475 480
Trp Phe Gly Phe Ser Gln Ser Asp Arg Met Lys Val His Ile Ala
485 490 495
Asp Gly Leu Asp Tyr Ile Ala Ser Leu Ala Gly Gly Gly Glu Ala
500 505 510
Arg Pro Cys Tyr Asp Val Ile Met Phe Asp Val Asp Ser Lys Asp
515 520 525
Pro Thr Leu Gly Met Ser Cys Pro Pro Pro A1a Phe Val Glu Gln
530 535 540
Ser Phe Leu Gln Lys Val Lys Ser Ile Leu Thr Pro G1u Gly Val
545 550 555
Phe Ile Leu Asn Leu Val Cys Arg Asp Leu Gly Leu Lys Asp Ser
560 565 . 570
Val Leu Ala Gly Leu Lys Ala Val Phe Pro Leu Leu Tyr Val Arg
575 580 585
Arg Ile Glu G1y Glu Val Asn Glu Ile Leu Phe Cys Gln Leu His
590 595 600
Pro Glu Gln Lys Leu Ala Thr Pro Glu Leu Leu Glu Thr Ala Gln
605 610 615
Ala Leu Glu Arg Thr Leu Arg Lys Pro Gly Arg Gly Trp Asp Asp
620 625 630
Thr Tyr Val Leu Ser Asp Met Leu Lys Thr Val Lys Ile Val
635 640
<210> 47
<211> 914
<212> PRT
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7766880CD1
<400> 47
Met Ile Gln Arg Ile Ser Cys Phe Ser Trp Ile Asp Val Phe Pro
1 5 10 15
Arg Gly Arg Leu Leu Ser Asp Glu Arg Asn Ile Leu Ser Asn Val
20 25 30
Asp Asp Ile Leu Ala Ala Thr Ala Ala Ala Cys Gly Val Thr Pro
35 40 45
Thr Asp Phe Ser Lys Ser Thr Ser Asn G1u Thr Met G1n Ala Val
50 55 60
Glu Asp Gly Asp Ser Lys Ser His Phe Gln Gln Ser Leu Asp Val
65 70 75
Arg His Val Thr Ser Asp Phe Asn Ser Met Thr Ala Thr Val Gly
80 85 90
49/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
Lys Pro Gln Asn Ile Asn Asp Thr Ser Leu Asn Gly Asn G1n Va1
95 100 105
Thr Val Asn Leu Ser Pro Val Pro Ala Leu G1n Ser Lys Met Thr
110 115 120
Leu Asp Gln Gln His Ile Glu Thr Pro Gly Gln Asn Ile Pro Thr
125 130 135
Lys Val Thr Ser A1a Val Val G1y Pro Ser His Glu Val Gln Glu
140 145 150
Gln Ser Ser Gly Pro Phe Lys Lys Gln Ser Ala Thr Asn Leu Glu
155 160 165
Ser Glu Glu Asp Ser Glu Ala Pro Val Asp Ser Thr Leu Asn Asn
170 175 180
Asn Arg Asn Gln Glu Phe Val Ser Ser Ser Arg Ser Ile Ser Gly
185 190 195
Glu Ser Ala Thr Ser Glu Ser Glu Phe Thr Leu Gly Gly Asp Asp
200 205 210
Ser Gly Val Ser Met Asn Pro Ala Arg Ser Ala Leu A1a Leu Leu
215 220 225
Ala Met Ala Gln Ser Gly Asp Ala Val Ser Val Lys Ile Glu Glu
230 235 240
Glu Asn Gln Asp Leu Met His Phe Asn Leu Gln Lys Lys Arg Ala
245 250 255
Lys Gly Lys Gly Gln Val Lys Glu Glu Asp Asn Ser Asn Gln Lys
260 265 270
Gln Leu Lys Arg Pro Ala Gln Gly Lys Arg Gln Asn Pro Arg Gly
275 280 285
Thr Asp Ile Tyr Leu Pro Tyr Thr Pro Pro Ser Ser Glu Ser Cys
290 295 300
His Asp Gly Tyr Gln His Gln Glu Lys Met Arg Gln Lys Ile Lys
305 310 315
Glu Val Glu Glu Lys Gln Pro Glu Val Lys Thr G1y Phe Ile Ala
320 325 330
Ser Phe Leu Asp Phe Leu Lys Ser Gly Pro Lys Gln G1n Phe Ser
335 340 345
Thr Leu Ala Val Arg Met Pro Asn Arg Thr Arg Arg Pro G1y Thr
350 355 360
Gln Met Val Arg Thr Phe Cys Pro Pro Pro Leu Pro Lys Pro Ser
365 370 375
Ser Thr Thr Pro Thr Pro Leu Val Ser Glu Thr Gly Gly Asn Ser
380 385 390
Pro Ser Asp Lys Val Asp Asn Glu Leu Lys Asn Leu G1u His Leu
395 400 405
Ser Ser Phe Ser Ser Asp Glu Asp Asp Pro Gly Tyr Ser Gln Asp
410 415 420
Ala Tyr Lys Ser Val Ser Thr Pro Leu Thr Thr Leu Asp Ala Thr
425 430 435
Ser Asp Lys Lys Lys Lys Thr G1u Ala Leu Gln Val Ala Thr Thr
440 445 450
Ser Pro Thr Ala Asn Thr Thr Gly Thr Ala Thr Thr Ser Ser Thr
455 460 465
Thr Val Gly Ala Val Lys Gln Glu Pro Leu His Ser Thr Ser Tyr
470 475 " 480
Ala Val Asn 21e Leu Glu Asn I1e Ser Ser Ser Glu Ser Ser Lys
485 490 495
Pro Ile Glu Leu Asp Gly Leu Pro Ser Asp Gln Phe Ala Lys Gly
500 505 510
G1n Asp Thr Val Ala Ile Glu Gly Phe Thr Asp G1u G1u Asp Thr
515 520 525
Glu Ser Gly Gly Glu Gly Gln Tyr Arg Glu Arg Asp Glu Phe Val
530 535 540
Val Lys Ile Glu Asp Ile Glu Thr Phe Lys Glu Ala Leu Lys Thr
545 550 555
Gly Lys Glu Pro Pro Ala Ile Trp Lys Val Gln Lys Ala Leu Leu
50/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
560 565 570
Gln Lys Phe Val Pro Glu Ile Arg Asp G1y G1n Arg Glu Phe Ala
575 580 585
Ala Thr Asn Ser Tyr Leu Gly Tyr Phe Gly Asp Ala Lys Ser Lys
590 595 600
Tyr Lys Arg Ile Tyr Val Lys Phe Ile Glu Asn Ala Asn Lys Lys
605 610 615
Glu Tyr Val Arg Val Cys Ser Lys Lys Pro Arg Asn Lys Pro Ser
620 625 630
Gln Thr Ile Arg Thr Val Gln Ala Lys Pro Ser Ser Ser Ser Lys
635 640 645
Thr Ser Asp Pro Leu Ala Ser Lys Thr Thr Thr Thr Lys Ala Pro
650 655 660
Ser Val Lys Pro Lys Val Lys Gln Pro Lys Val Lys Ala Glu Pro
665 670 675
Pro Pro Lys Lys Arg Lys Lys Trp Lys Glu Glu Phe Ser Ser Ser
680 685 690
Gln Ser Asp Ser Ser Pro Glu Tle His Thr Ser Ser Ser Asp Asp
695 700 705
G1u Glu Phe Glu Pro Pro Ala Pro Phe Va1 Thr Arg Phe Leu Asn
710 715 720
Thr Arg Ala Met Lys Glu Thr Phe Lys Ser Tyr Met Glu Leu Leu
725 730 735
Val Ser Ile A1a Leu Asp Pro Asp Thr Met Gln Ala Leu Glu Lys
740 745 750
Ser Asn Asp Glu Leu Leu Leu Pro His Met Lys Lys Ile Asp Gly
755 760 765
Met Leu Asn Asp Asn Arg Lys Arg Leu Leu Leu Asn Leu His Leu
770 775 780
Asp Gln Ser Phe Lys Asn A1a Leu Glu Ser Phe Pro Glu Leu Thr
785 790 795
Ile Ile Thr Arg Asp Ser Lys Ala Lys Ser Gly Gly Thr Ala Ile
800 805 810
Ser Lys Ile Lys Met Asn Gly Lys Ala Tyr Asn Lys Lys Thr Leu
815 820 825
Arg Thr Ser Lys Thr Thr Thr Lys Ser Ala Gln Glu Phe Ala Va1
830 835 840
Asp Pro Glu Lys Ile Gln Leu Tyr Ser Leu Tyr His Ser Leu His
845 850 855
His Tyr Lys Tyr His Val Tyr Leu Ile Cys Lys Asp Glu Ile Ser
860 865 870
Ser Val Gln Lys Lys Asn Glu Asp Leu Gly Gln G1u Glu Ile Val
875 880 885
Gln Leu Cys Met Lys Asn Val Lys Trp Val Glu Asp Leu Phe Glu
890 895 900
Lys Phe Gly Glu Leu Leu Asn His Val Gln Gln Lys Cys Ser
905 910
<210> 48
<211> 148
<212> PRT
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 90089609CD1
<400> 48
Met His Gly A1a Arg Leu Ser Asp Lys Leu Trp Arg Val Met Glu
1 5 10 15
Leu Trp Leu Gly Arg Ala Gly Arg Leu Gly Arg Lys Gly Leu Arg
20 25 30
Gly Arg Arg Ala G1y Arg Ala Gly His Thr Gly Leu His Gly Gly
51/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
35 40 45
Ala Ala His Leu Arg His Pro Gly Val Pro Ala Ala Val Ala Phe
50 55 60
Leu His Leu Pro Gln Asp Ile Ser Val Gln G1n Thr Leu Asp Leu
65 70 75
Val Pro Val Leu Glu Gly Leu Ala Val Ser Thr His Pro A1a Leu
80 85 90
Gln Leu Arg Arg Leu Ala Cys Gly Gly Trp Gln Thr Arg Pro Gln
95 100 105
Ala Ala Ala Pro Pro Leu Trp Asp Phe Ser Gln Gly Arg Arg A1a
110 115 120
Asp Gly Val Ser Val Thr Ala Ser Tyr Arg Ala Leu Gly Gly Gly
125 130 135
Pro Cys Ser Arg Ser Arg Pro Leu Pro Arg Pro His Thr
140 145
<210> 49
<211> 882
<212> DNA
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No. 1629602CB1
<400> 49
cagcggccgg ccgcggggcc tttttgtcct gagggccaga gaaagggaga agggggtggg 60
gggacagcca cgtggccgca ggaggattta caacattttc tttcgccatc gatgttatcg 120
caaaatgtgt gagagaagcg gctgcgcagc ccggacggga gcgtgagggt gcgggccagg 180
taagcagccc cggcggtttc gccgcatacg ggactgcggg gcgaccgcgg gcaccagcca 240
cgcgcagcgg ctccgcgggg tctcggccgg gtccgcgctc tgaaggatct cgagagccat 300
ggatggtgca ggcgggaccc tcgagctgca gcatctccgg tgacccgggg ttgccgagga 360
ggtggagacc agcacaggtg gtccggcccg ggcgcctccg aatccggggg tggtcaagac 420
ggatccccaa ggctgaggtc ggcagtcccg gggactcgca gctgttgagc ctgtggagac 480
gcggccccgt gaccgaggca cccttcagca acccgggggc agcgttttcc ccctaccgga 540
aatctgatgg gcttatgaca tcatggctgg ctgctgagcg atgaagtgga tgccacaaag 600
aaatccgaca tatcagatag attctgaaat cggtttccct ccagctgtag taacaggcgt 660
gaagtcagga gaatttgagc tttgtttaaa aaataaataa ataaataaat aaaccataac 720
aaagtcttgc cctgtattaa atgcaatttt Cttaaaaaca agcaaacctt ttggacatca 780
ttttatttta atagaaatgc tgagttttat gaaactaaag tggctaataa atcagacctg 840
aagctttgtg tgagtgttcc aaaaaaaaaa aaaaaaaaaa tg 882
<210> 50
<211> 2489
<212> DNA
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 2100360CB1
<400> 50
gggcactctg gtgtacagcc agtccccgcc gcggaggtgc cggtggagcc tgggaccggg 60
cgagtctccg ccccgctttt gcagctaggg gtgtgtttca ggggggattg gggcaagcca 120
agcaggcgag gacctgggcc tgtgccgctt tgcctacccc tcatccctcg gcaccaaggc 180
tacttgagcc ccagggtgtt ttttccttgt tcccgccacc tcctggtccc tggcccaaca 240
tgatactgac caaagctcag tacgacgaga tagcccagtg cctagtgtct gtgccgccta 300
ccaggcagag cctgaggaag ctgaagcaga ggtttcccag tcaatcgcag gccactctgc 360
tgagcatctt ctcccaggag taccagaaac acattaaaag aacacatgcc aaacatcata 420
cttcggaagc aattgaaagt tattaccaga ggtacctgaa tggagtggtg aaaaatggag 480
ctgccccagt gctcctggac ctggccaatg aggtggacta tgcgccctca ttaatggctc 540
ggcttatact ggagaggttt ctacaggaac acgaggaaac tccaccctcc aagtctatta 600
taaatagtat gctacgggac ccttctcaga ttccagatgg agttctagca aatcaggtct 660
52/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
atcagtgcat tgtgaacgac tgctgttacg gaccactagt ggactgcatc aagcatgcca 720
ttggtcatga gcatgaggtc ctgctgagag acttgcttct agagaaaaac ctgtccttcc 780
tagatgaaga tcagcttcgt gcaaagggtt atgacaaaac accagacttc attttacaag 840
taccagttgc tgtagaaggg cacataattc actggattga aagcaaagcc tcatttggtg 900
atgaatgtag ccaccacgcc tacctgcatg accagttctg gagctactgg aatagatttg 960
ggccaggctt agtcatctat tggtatggat ttatccagga gctggactgc aaccgggaaa 1020
ggggcatcct gctcaaagcc tgtttcccca cgaacattgt caccttatgc cacagcatag 1080
cttgaccctg aagatcctgg aagagaagct gggaggaaaa gagacccagc attgcattac 1140
catcgtggaa taatctagcg caaacctagg aaagctgaag ccacaaagtc caaagccacc 1200
tttgtactca cctgcagagc tccagaagac cttgatggca gcctgcctat gctgtgtgtt 1260
tgctatattc aatctttacg gcttcctgac ttctgtgaca gtaagccaag tgcaaaaata 1320
cacttgatga gaatttcctc ttttaataat gttatttgaa caccacatat tttagattta 1380
tcttatttga aagtattagt tccattgtgc ctggaaacca cactccttta gattgggggc 1440
cgagaggcga caacccaaca ttgaggagag tttattttta aacatggcta gttgtcagta 1500
tgtacgtgag ctagtatttt tatgagtcga gttttttaaa ggcacattct gtatactgct 1560
tagtatatgc attttatacc atgtaattat aaaacactcg agtaagttca gcattagaaa 1620
tgtttagctt tgtatgaact gagtgtgcca gaaataaacc tggagcaatt tttaaataag 1680
caaaataaag gagatttttc tatttgttca ctttaattta ttcacttttg tgtactttta 1740
tgtactgcaa atcagatttc agtctaaagc gaaacatcag taagttaata ataaacctat 1800
ctttcgggaa gttgaatatt aatctgtacc caaaacgtat ttagtaaaat atttgccccc 1860
gccaccctgc catgctgaca taacaacttt tataatgttg aatagatgat atgggaaata 1920
ctaataacaa caatgtaatt tttgcagaca gctttaactt atatacattg cttgattttt 1980
ttcaaaagac taaatatgtc atttatactt tgtttatttt ctaccaaaga aggtttgtaa 2040
aaatatgcct gctgcttttc cttttgaagg acacaaacct ggtcccaaca tgtgtggatt 2100
ttaactctga gtggggtgca ttaaatcaaa agagagaggc agaagatgaa atgctaaaga 2160
agggtcaggc aaacttctgt ttcagtataa aattcatcat gcaggcttct gagtgaaata 2220
gaatgatttg aaaccactac tgtattgcct ggatacacac acacacacac acacacttta 2280
tacaaaaatg ttaaaagcag gtttcctggc atgttctaaa ctgttttttc tttaggaata 2340
aattacattt atctgcacag atgttgaaaa tcctgttaaa cccttgtcaa ggatttgttt 2400.
attttacatt aaacaaattt attatgatga acgtgaacaa ataaattaaa aaataaaaaa 2460
ggtaaaaaaa aaaaaaaaaa aaaattcgg 2489
<210> 51
<211> 1115
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Tncyte ID No: 5166833CB1
<400> 51
ctcgaacttg gtcggggcgc ggatcccgag agggaaagtc ataacaaccg cacgagggag 60
ttcgactggc gaactggaag gccacgcctc ctcccgcctg ccccctcagc cctgtggctg 120
ggggcagagc tcagactgtc ttctgaagat tgatgtctat ttccttgagc tctttaattt 180
tgttgccaat ttggataaac atggcacaaa tccagcaggg aggtccagat gaaaaagaaa 240
agactaccgc actgaaagat ttattatcta ggatagattt ggatgaacta atgaaaaaag 300
atgaaccgcc tcttgatttt cctgataccc tggaaggatt tgaatatgct tttaatgaaa 360
agggacagtt aagacacata aaaactgggg aaccatttgt ttttaactac cgggaagatt 420
tacacagatg gaaccagaaa agatacgagg ctctaggaga gatcatcacg aagtatgtat 480
atgagctcct ggaaaaggat tgtaatttga aaaaagtatc tattccagta gatgccactg 540
agagtgaacc aaagagtttt atctttatga gtgaggatgc tttgacaaat ccacagaaac 600
tgatggtttt aattcatggt agtggtgttg tcagggcagg gcagtgggct agaagactta 660
ttataaatga agatctggac agtggcacac agataccgtt tattaaaaga gctgtggctg 720
aaggatatgg agtaatagta ctaaatccca atgaaaacta tattgaagta gaaaagccga 780
agatacacgt acagtcatca tctgatagtt cagatgaacc agcagaaaaa cgggaaagaa 840
aagataaagt ttctaaagaa acaaagaagc gacgtgattt ctatgagaag tatcgtaacc 900
cccaaaaaaa aaaagaaatg atgcaattgt atatcagagt gagtgagatc actactttcc 960
tttactattt tctttacctt gtatatattt tattatatgt agattgtttt gtttttcttc 1020
aagaatatta atttctttat ttgtcatcat ttatttccca tggtcgtcta cttggattaa 1080
atgggttttt aaattcaaaa aaaaaaaaaa aaaaa 1115
<210> 52
53/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<211> 2434
<212> DNA
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte TD No: 7494963CB1
<400> 52
cttgcaatgt tttgttaagg aaatggagtt gtttgtccta tagaattttc ttttttcttt 60
ttttcttttt gaggtggcgt ctcgcactgt cacccaggtt ggagtgcaat ggcgtgatct 120
tagctcattg caacgtccgc ctcctgggtt caagcgattc tcctgcctca ttccccaagt 180
agctgggatt tcaggtaccc gccaccacgc ccagctaatt tttgtatttt tagtagaggc 240
agggtttcat catgttggcc aggctggtct catactcctg accttaggtg atctgc,ccgc 300
ctcggcctcc caaagtgctg ggattacagg ggtgagccac cgcgcccagt cagaattttc 360
tagatctgga ttttgctgag tgcatctcct gatgtagttt aacatgttct tctctcctat 420
ttatttttct ataaatggtg gttggatcca gaagcatgat caggttgagg gtcagttgtc 480
ttggttttgt gtttggtaaa cctcaaccat ttgctctttc tttgaaagct gcaccaaagc 540
cagctggagc ttcaggaggt gcgtctctcc tgccgacagc tgcaggtgaa ggtggaagaa 600
ctcactgagg agaggagtct gcagagctct gccgccacca gcacatccct cctgtcagag 660
atcgagcaga gcatggaggc tgaggagctg gagcaggagc gagagcagct gagactgcag 720
ctctgggaag cctactgcca ggttcgctat ctgtgctcac accttcgagg caatgacagt 780
gctgactcag ccgtctccac ggactcctcc atggacgagt cttcagaaac ctcgtccgcc 840
aaggatgtgc cagccggcag cttgcgcact gccctcaatg agctcaagag actgatacag 900
agcattgtgg atggcatgga gcccacgggc tcccggagac ttgatgatga Ctccttagaa 960
gaacagataa ggcagaccag tgaggactcg agagccctaa gggagctcat ggagggagag 1020
aggggtaaac tgaggcaaag cctagaagag ctgcagcgac tccacagtca ggtgacactg 1080
ctgagtgtgg agatgactgc cctaaaagag gagagagacc gactcagagt cacttctgag 114 0
gacaaggagc caaaggagca gcttcagaag gccatcaggg accgcgacga ggccattgca 1200
aagaagaatg ctgtggagct ggaacttgcc aagtgcagaa tggatatgat gtctctgaac 1260
agccagttgc tggatgccat tcagcagaaa ctgaacctct cgcagcagct ggaagcttgg 1320
caggatgaca tgcacagggt cattgaccgg cagctgatgg acacgcacct gaaagaacgg 1380
agccagccgg ctgctgccct ctgcaggggc cacagcgctg ggcgggggga tgagcccagc 1440
atcgctgaag gcaaacgact cttctcattc ttcaggaaaa tttaagttgg gaggagtcag 1500
gccaccaaag atgggtggac tggaggcagc tggaaaggcg gtgcaggcaa ggcctcccct 1560
gcagcttgca cctcagcagc tgccctgccc ctcatgctag ggccccatgg gtccgggagg 1620
gcctgctccc tttcgtcggt ggggatggag acctagaggt gggggcctgc cttggccact 1680
gaaggcttcc cttggcccac cgcctggcca agcccacgcc tgggcttctc caggaccacg 1740
tgcttgagca gggttaggcc acctcccaga ggggcccctt ggtgttgggc tttgcagctC 1800
acacccaaca gatcgcagcc cacccccagg cactgctgcc tccttgattt tagcaaatgg 1860
ggaacagaag gaatggaggc ccttctctgc atgcctcagg aggcctgagc cccaggggcc 1920
tagacctgtg ggggcagcgg gccaggcctg agcctccatt ccttccccag cccctggccc 1980
agggtcaaag gagagatggc agcccctccc ccgcatgcat gcacctcagc tggcaggagg 2040
ccaagcctct ggccgcaggg tctaagagcc ggggcttacc caagctcagc tgaggccacc 2100
cgagccccag ggaggaagaa ggccctgtcc ccctgtcgcc actgctctcc ctcccagcct 2160
tcagtctctg ccccttagca gggcctggcc aggcagagtg ttatcaccag tcatctgcag 2220
gctttagcca tccagccctt tcccctgctc agggctgggg ttggacgggg tctcctcctc 2280
ccacagctcc ctcctccacc cctcacatac atacataatt tcttggccta gccaaacaag 2340
tccaggccac tgaatggcac cagaggagtc tgtggtcagc caccccacct tgagggcagc 2400
acaggcacca cggggtcgag gggaggggga ggct 2434
<210> 53
<211> 3492
<212> DNA
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7644881CB1
<400> 53
gctactttag actttttcat ggttatcaat ctgtacaaag aatcaccaaa ctgataaagc 60
aggaaccaga gggcaaatca cgctgccaag acaactgtgt aattcgctcg aaaaagaaac 120
54/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
gatggagtct cgctctgtca cccaggctgg agtatagtgg tgtgatcttg gctcactgca 180
acctctggcc tcccaggttc aagcgattct cctgcctcag cctctcgagt agctggaatt 240
acaggtctgg cagaaggaac agtatcaact gactgagtag gtctcattgg cagttgtgat 300
tcagagacct agaaagctga acccacggct ggcaagaaga ggatggtttg tgggacctgg 360
gctgatgtct gatgaaattt taagccccag ctatagctac tacaaagaaa agtggctgat 420
gataagcatg taactcaaaa agacaatgta tataaaaata tgcaagaatc acaggaaacc 480
cacatatcca accacctaga tgaagttgtt gctgctgtta gcatcactca tagaaagaag 540
ttccaaaaca agctgcttca gacagcacta ttccagcctc ctcgagagaa actccacctc 600
tgtgaagaga aagcaaagtc ctattccaac agtcatgagt acaaacaggc cgtccatgag 660
cttgtgcgtt gcgtagcact gacaagaatt tgctatggag actcacattg gaaactagca 720
gaggcacatg ttaatctggc tcaaggctac ctccagctga aaggactgtc actgcaagca 780
aaacaacatg cagaaaaagc cagacaaatc ctcgccaact ccattgtgcc tccctatagt 840
gagaatacag atgttttcaa gttttccatt gagcttttcc ataccatggg cagagcttta 900
ctctcccttc aaaaatttaa ggaagctgca gagaatttga caaaagcaga gagactttca 960
aaggagctgc tacaatgtgg aagaattata aaggaagaat ggatagaaat tgaagcacgg 1020
atcagattat catttgcaca ggtgtatcaa ggtcagaaga agtcaaaaga agctttgtcc 1080
cactatcaag cagctttgga atatgttgag atcagtaaag gtgaaacaag tcgtgagtgt 1140
gtacccatat tgagagaatt agcaggtgta gagcaagccc tgggactcca cgatgtatcc 1200
atcaaccact tcctccaggc acatctcatc atcctgagta gaagcccctc tcaagtggag 1260
gcagcagact cggcacacat cgtcgcccat gctgctgtcg cttcagggag acacgagcac 1320
catgatgtag ctgagcagta ttttcaagag agcatggctc atcttaagga ttctgaaggg 1380
atgggaagaa ccaaatttct ttcaattcaa gatgaatttt gccattttct acaaatgact 1440
ggacaaaaag agagagcaac ctcgatcctg agagagtccc tggaagccaa agtggaagca 1500
tttggcgatt tcagtcccga ggtggcagag acataccggc tcctgggagg agcagacctg 1560
gcgcagggga accacagtgg ggcccgcaag aaactgaaga agtgtctcca gatccagacc 1620
ctcttatatg gaccgcagga caaaaggact ctggccaccc agcaggccat gggcatgctg 1680
tccacggccc ccaaggttgc ttcgaagcca aggcaggcat caaaagccaa agtggccttc 1740
tgcaccagca tccctcagga caccctgctg gggaaggccc ggcccggcac aacagcagac 1800
tgaggccccc accctgaaaa agcctaggaC attcctgggc actgtcattt agggtgctgt 1860
acaaatcacc tccgcctaga aaatggaatt caacagtcag gatacagatt tccaaggcca 1920
actgttggcc ccaacatgca acagtgagac cataagcctc ccgtgggcca cattttgaca 1980
gtggatgccc ttcagggtga tatatgctat aaagcagttt tctatcacct aagtggtttt 2040
tcttgccaac aagaattttt acccatcagc actactgtgg ctgaaaaact tctcttcaac 2100
agttcagtgc gccctgtgca ggagtcagcc cggcatctgc ttgtacacac agctccttgc 2160
ataggtgtgg agttagatct ggacagtgaa cttcaggaag tcctttctta taggaggcta 2220
atagggattg agaataacat gagaagaaaa cgctaataaa gggaaacctg aacacgctgc 2280
tgtcagcatg tgttttcaaa gtgcagcctg cctcagagtt cttcggagcc tgaaaagggg 2340
tttgagaaag agcccagtag gaggggcagg aggccgacac acctgacttg gcctggggcc 2400
caggaggcag gtgtaaggga gtgaaaagaa aggctagccg gaggctgcgg ggggaagacc 2460
gcagactccc tgctgcttcg catccctcct gtggcctcca ctgcaggcag gacaaacctg 2520
gatgccacct ggagctgctt cctgagttgg cacactatcg tgtacacagc agtcttcagc 2580
cccctggaag gaggccatag tcgtgtgagg atggcaaagt cgaacaggaa gctttgagtg 2640
ccttcctcca cgatgtcaac gaggagatcc agtgccagat cgaggtggat ggaacaccca 2700
ggggtagggg tgcaggtgtg ggcagtgatg tcccttcccc tccctcccct ggtcccacag 2760
actgtggcca tgaggctgca ggctggtgct atgacagcag attgcagcac agggccctcc 2820
cctccagccc ccagtgggac atcaaaacca ccctggggcc atttgtgcag ggcaccacct 2880
ccagtattga tggggaaaat aaactcagta gagccacgac agggtggaga gaagcaggga 2940
ccattgtctt cctcaggagc gtgacagctg accccacaga ccatgcttgc tggtacacac 3000
tggtcccaga cccaggcctg tcggacatca gcagtgtgct aaaaacgtgt aagatgtggt 3060
cactactcac cgtgtgtcct atctagttga catgggtgga gtcagctaag gggtgaatgt 3120
tcatatgctc ccaattcacg ttgaagccct aatccccaaa gggatggtat tgggggtggg 3180
gtcttggaga ggtgattagg ttatgagggt ggagccctga tgaatgggat tagtgcttta 3240
taaggagaga caccagagag atgatctctc tctccaccat gtgaggacac agtgagaaga 3300
cagccgtctg caggcccgga agagagccct caccaggaaa tgaaactgtt ggcaccttga 3360
gacttcccag cttccagaac tgtaagaaat aaatgtttgt tgtttaagcc tttcaggcta 3420
cggctttctg ttacagcagc ctgaactgag agtccatgcc gagtttttga aataaatgtg 3480
aattctgatg tt 3492
<210> 54
<211> 3141
<212> DNA
<213> Homo sapiens
55/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<220>
<221> misc_feature
<223> Incyte ID No: 3790383CB1
<400> 54
tccacacccg ctggctggct gatgtttgat tctgtaacta taccacgccc agaattctct 60
caaaagggaa taaaacacag gtcaaattcc tcacccacac actccacagt tcaacccctg 120
ccagggaaac caaaagcagg aaaggatctc cagcggcgcc attctcattt ccggtcccag 180
caccccgcct ccatgacgtc aacgcgccgc gccaccgggc tgcgtcatct cggcgcgccg 240
ctgccagggc tgtacacctg ctggctgcca tggctgaggt gggccgtacc gggatcagct 300
acccaggcgc gcttctccca cagggcttct gggctgcggt cgaagtgtgg ctggagaggc 360
cgcaggtggc aaacaaacgg ctttgcggcg cccgcctgga ggcccgctgg agcgccgccc 420
tgccctgcgc ggaggcccgc ggccccggga ctagcgcagg ctcggagcag aaggagcggg 480
gtccgggacc cggccagggt tcccccggag ggggcccggg tcccaggtcg ctatcaggac 540
ccgagcaggg cacggcatgt tgcgaacttg aggaggccca gggccagtgc cagcaagagg 600
aggcacagag ggaagccgcc tcagtgcccc tgagggactc cgggcacccc ggccatgctg 660
aaggaaggga gggcgacttc cccgccgcag atctggattc gctttgggag gatttctccc 720
aaagtctcgc ccgtggcaat tcggagttgc tggccttcct caccagctcc ggggcgggat 780
cgcagccaga ggcgcagcgt gagctcgacg tggttctcag aaccgtcatc ccgaaaacta 840
gcccacattg cccccttaca actcccagga gggaaatagt cgtgcaagat gtcctcaatg 900
gaaccataac gtttttgcct ttggaagaag atgatgaggg gaacctaaag gttaagatga 960
gcaatgtgta tcaaattcag ctcagtcata gcaaagaaga atggttcata tctgttttaa 1020
ttttctgtcc agaaagatgg cattcagatg gaatcgtgta tcccaaaccc acgtggcttg 1080
gagaagagtt gctggccaag ttggccaagt ggtctgtaga gaacaagaag agtgacttta 1140
aaagcaccct ttccctcatc tccattatga agtatagcaa ggcttaccag gaacttaaag 1200
agaagtataa ggaaatggtt aaggtgtggc ctgaagtcaC tgatcctgag aagttcgtgt 1260
atgaagatgt ggctatcgca gcatacctgc tgattctatg ggaagaagaa agggctgaga 1320
ggggactaac tgccaggcag tcctttgtgg acctgggatg tggaaatggc ctcctggtcc 1380
acatcctgag cagtgagggg catccaggca gagggattga tgtccgaaga agaaaaatct 1440
gggacatgta tggaccacaa actcagttag aggaagatgc aatcacaccc aatgataaga 2500
cccttttccc tgatgttgat tggttaatcg gtaaccattc tgatgaactc acaccatgga 1560
tacctgtcat tgcagccagg tcttcctaca attgccgctt ctttgtcctc ccctgctgct 1620
tctttgactt cattggaaga tactcccgga ggcagagtaa gaagactcag taccgggaat 1680
accttgactt cattaaagaa gtgggcttca cctgtgggtt tcacgtggac gaagactgcc 2740
tcaggattcc ttcaaccaaa agagtctgtc tcgttggaaa atccagaaca tacccttcct 1800
ccagagaagc ttccgtggat gaaaagagga ctcagtacat taagagcagg cggggctgcc 1860
ctgtaagccc acctggctgg gagctttccc cttctccacg ctgggttgct gctggcagtg 1920
ctggtcactg tgacggtcag caagctctgg acgccagggt cgggtgtgta accagggcct 2980
gggccgctga gcatggagca gggccccagg ctgaaggacc ctggctacct ggatttcatc 2040
ccagagaaaa ggctgagcgt gtgaggaact gtgccgccct gccacgagat tttattgacc 2100
aagtggtttt gcaagtagcg aatttactgt taggtggaaa gcaattaaac acaagaagtt 2160
ctcgaaatgg gagtttgaag acctggaatg ggggagagag cctatctctg gcagaagtag 2220
ccaacgagct ggacacggag accctgcgga ggctgaagcg ggagtgtggg ggcctgcaga 2280
cgctgctccg gaacagccac caggtgttcc aagttgtgaa tgggagagtt cacatccgcg 2340
actggcgaga ggagacactg tggaagacaa agcaaccgga agcgaaacag agactgctct 2400
ctgaagcctg caaaacccgc ctctgctggt tcttcatgca tcaccctgat ggctgcgctc 2460
tgtccacgga ctgctgcccg tttgcccatg ggcctgcgga gctgcggcca ccccggacca 2520
ccccgaggaa gaagatttca tgagctgcat ccttgccagc cgaggcctgg ttggggaggc 2580
caaaccaagg agagcttccc cagcagtcgt cagtgctgtg gtctctgctc tggctgtgtt 2640
tcagcccacc tcctcccagc tttctccaca tcctcacagt gatgaaccgt atttcataaa 2700
catcacacgc cagagaagcc acagttactc ggaagccccc agctgactgc ctggcttgtt 2760
tcagatgcag ccgcttgaaa cgtgcgcagc atcttcatat cataaagatt gtgcacggat 2820
Ccttacaatg tctcctgggg gagagcggct gaggctgcct tgcacaggcc cttcccaggg 2880
cgctgtccga cgcctgcccc accatgtcca catctgtgaa gaggatgggg ctcctcgaga 2940
agtaagaccg tatctgccag cgtttctcac cacactggag agcagctgct ctggagcagg 3000
gatccaccag attggtattt ttaaaaaagg tgtcaggctt gctatgttga ggttgttttt 3060
agagttacag agaataaaaa cactcataat ttcctgaaaa aaaaaaaaaa aaaaaaaaaa 3120
aaaaaaaaaa aaaaaaaaag g 3141
<210> 55
<212> 3491
<212> DNA
<213> Homo sapiens
56/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<220>
<221> misc_feature
<223> Incyte ID No: 3846110CB1
<400> 55
cacgaggcgc agctatacaa ctgggccgca ccagaagtga tcttacagaa ggcagccaca 60
gtgaaatcag acatctacag cttttctatg atcatgcagg agattttaac agatgacata 120
ccctggaagg gcttatatgg ctcagttgtt aaaaaagccg tagtctcggg gaattattta 180
gaagctgatg tcaggcttcc gaaaccttac tatgatattg ttaagtcagg catccacgtc 240
aagcataaag accgaactat gaaccttcaa gatatccggt atattctgaa gaatgactta 300
aaggatttta ctggagccca gagaactcaa ccaaccgaga gccccagagt gcagagatac 360
ggactccatc ccgatgtcaa tgtctatcta ggactgactt cagaacaccc cagagagaca 420
cctgacatgg aaatcataga actaaaggaa atgggcagtc aacctcattc accaagggtt 480
cactctttat tcactgaggg gacactagat cctcaggccc cagatccatg tctgatggcc 540
agggagactc agaatcaaga tgctccttgc cctgctccat ttatggcaga agaggccagc 600
agccccagca caggtcagcc aagcctctgc agtttcgaaa tcaacgagat ctactcaggc 660
tgcttgattt tggaagatga catagaagag cctccaggag ctgcttcatc tttggaggca 720
gacggaccta accaggtaga tgaactgaaa tccatggaag aagagctgga taagatggag 780
agagaggcgt gttgttttgg cagtgaggat gagagctctt caaaagctga gacagagtac 840
tcttttgatg actgggactg gcaaaacggt tcactcagtt cactcagcct tcctgagtca 900
accagagaag ccaagagcaa tttgaacaac atgtccacga ctgaggagta tctcatcagt 960
aagtgtgtgc tggatctaaa gattatgcag acaataatgc acgagaatga tgataggctg 1020
aggaatatcg agcagatatt agatgaagtc gagatgaaac agaaggaaca ggaagagcgc 1080
atgtctttat gggccacttc aagagagttt acaaatgcct acaagttacc tctggccgtg.1140
ggccctccat ctttaaacta tattcctcct gtcctacagc tttcaggggg tcagaagcca 1200
gacaccagtg gcaactaccc aaccctacca agatttccaa gaatgctgcc gactctttgt 1260
gaccctggaa aacagaacaC agatgaacaa tttcagtgca ctcaaggagc caaggacagt 1320
ttggaaacaa gcaggatcca aaataccagt agccagggaa gacctagaga gtccactgcc 1380
caagccaaag ccacacagtt taatagtgca ctcttcactc tgtcaagcca ccggcaggga 2440
ccttctgcat cacccagctg tcactgggac tctaccagga tgagtgtgga acctgtttct 1500
tctgaaatct ataatgcaga gtccagaaat aaagatgatg gaaaggtaca cttaaaatgg 1560
aaaatggagg tgaaagaaat ggcaaagaaa gcagctactg gacagctcac agtacctcct 1620
tggcatcctc agagtagtct gactttagag agcgaggctg aaaatgagcc cgacgccctg 1680
ctgcagcccc ccattaggag cccagaaaac acggattggc agcgagttat tgagtatcat 1740
agggaaaatg atgagcccag aggaaatggc aagtttgaca agacgggcaa caatgactgt 1800
gacagtgacc agcatggcag acagcccagg cttggaagct tcaccagtat caggcaccca 1860
tctcccagac aaaaggagca accagagcat agtgaagcct tccaagcaag ttctgacaca 192'0
ttggtggctg tagagaaatc ttacagtacc tcgagtccca tagaagagga ctttgaagga 1980
atacaaggtg catttgccca acctcaagtc tctggtgagg aaaagttcca aatgagaaaa 2040
attcttggaa agaatgctga gattttgccc aggtctcaat ttcaacctgt acgaagtact 2100
gaagatgaac aagaagagac atcaaaggag tcaccaaagg aactgaaaga gaaagacata 2160
tcattgacgg atattcaaga cctgtctagt atctcctatg aaccagacag ctcttttaag 2220
gaagcttcat gcaaaacacc caaaataaac catgcaccta ccagtgtcag cactccactc 2280
agcccagggt ccgtttcttc agctgccagt cagtataaag actgccttga aagtatcaca 2340
tttcaggtta agacagagtt tgcctcttgc tggaacagtc aagaatttat tcaaactttg 2400
tctgatgact ttataagtgt ccgagagaga gcaaaggaac tggattctct ccttacttcc 2460
tctgaaactc ccccttcaag actgactggt cttaaaagat tgtcttcatt tattggggct 2520
ggatccccca gccttgttaa ggcatgtgac tcatcaccac cccatgccac ccagagaagg 2580
agcctgccta aagtagaagc cttctcacag catcacattg atgagctgcc accaccatct 2640
caggagctac ttgatgacat tgagctcttg aaacagcagc agggctcatc cacggtgttg 2700
catgagaaca cagcaagtga tggaggaggc actgcaaacg atcaaaggca cttagaagaa 2760
caagaaactg acagtaaaaa agaagatagt agtatgcttt tgtccaaaga aactgaagat 2820
cttggagagg acacagagag agctcactct actctggatg aggacctgga aagatggctg 2880
cagccacctg aggagagcgt ggagctacaa gaccttccca agggctctga aagggagaca 2940
aatatcaaag atcaaaaagt tggtgaagag aaaagaaaaa gggaagatag cattacacca 3000
gagagaagga aatcagaggg tgttctaggg acttctgaag aagatgaact aaaatcctgt 3060
ttttggaagc gactaggttg gtccgaatca tccaggataa tcgtgctgga tcagagtgac 3120
ttgtcagact gattggaatt ggatcataga cggactcctg gcctgagttt gagtgtcctg 3180
gttgtaagct cctttcttct ctttctgctt cagttgctgt cagggcagca gttccagttc 3240
tgtaagtctc actttgttca gctgccacaa tagacatcat cgtttggccc tctctgttag 3300
cagcacattc aaccatttgt tttcagtcag atttctgaaa agtgagaggt agttttgata 3360
gtaaaaattt ttggttgtgc ctagaatggc tttggttttg ttgatgttaa ttttcaaaaa 3420
ctttaactct tgttatataa taaaatgttt aattttaata acagaaaaaa gggggtccac 3480
57/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
tagtttagag a 3491
<210> 56
<211> 4312
<212> DNA
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 1878279CB1
<400> 56
tcttatgcca tgcctctagt gtgccacatt tatgttcacg atctcattta attcttgcca 60
caaccctatc aggaaggtgg gagtcaatca ttttgacaag tctcctgaaa ggaacagcta 120
gcaggaactg aaaccttttt ccatttggtc tcgtggcaaa ggcagagatt gctccagcag 180
ctccacacaa aatgatgtgc tcacgggtgc cctctgaaca gtcttctggt acctctctct 240
tgcctaaaga cggtgcccca ttttcttggg attccttgga tgaggatgga ttggatgact 300
ccttgctgga gctgtcagag ggagaagaag atgatggtga tgtaaattac acagaggaag 360
agattgatgc actgttgaag gaagatgacc catcatatga gcagtcttct ggggaagatg 420
atggtgggca tgttgagaag ggagaaagag ggagtcaaat tctacttgat actccccgag 480
agaaaaattc atcgtacagc ctgggaccag tagctgagac tcctgacctc ttcaaactac 540
ctcagctaag tacatcaagt ggtcatggac cagctcatac taaaccatta aacagacgct 600
ctgtactaga aaagaatctt ataaaagtaa ctgttgcacc atttaatcca acagtttgtg 660
atgctctgct tgataaggac gagactgatt cgtccaaaga tactgaaaaa ctctcttccc 720
ttggagaaga gatgagagaa gatggtctta gcccaaatga aagcaaactt tgtactgaat 780
ctgaagggat cagccccaat aactctgcct ggaatgggcc ccagctctct tcttcaaaca 840
ataactttca acagactgtc tctgataaaa atatgcctga cagtgagaac cctacgtctg 900
tattctctcg gatctcagac cattcagaga ctcctaatat ggagttatcc tgcagaaatg 960
gtggttcaca caagtcaagt tgtgaaatga gatctctggt tgtttccacc tcatcaaaca 1020
aacaggatgt tcttaacaag gattctggga agatgaaagg ccatgagaga agactaggca 1080
aagtcattcc tgttctacaa actaagacca ggactaatgt tccgacgttt tcacagtcaa 1140
atctagaaca gcagaagcag ctttatctca ggagtgtcat tgctcatata gaagacccag 1200
aggacactaa ccaaggatga atccctagaa gtggaattgc tgggtatctc gggggagctt 1260
tgtgccttga tggatcaagt tcatcatatg cagcactcaa aatggcagca tccttcggac 1320
ctcaccacgc gaaactacgc ccgccgacag aaacatctgc aaagatacag tctgactcag 1380
tgggttgaca ggaacatgcg aagccaccat cggttccagc gtctcccaga Cttctcgtac 1440
agttaatttg tgtcatccca tcagcaatga aggtccctat ccagggtcct gcttggagca 1500
gcatttcatg ttcttttgct gttttgtgct ttgccgattt tggattttat ttttcacaaa 1560
atttttattt aaaaaactcg tcaccttttg gaaatgccca ttgccgactt gaattttttt 1620
gtatgaagtc cctcctgatt ttgtgtgtgt gtgtctgtgt ttaagcaagc gttcggttgg 1680
tatagttttt ttttgttttt ttaatttaaa ttgaaggtag ctgcctcctg aaagccagca 1740
ttaagccaga acacccaggt tcaagcaaaa gacccacctc tctgcagagg caaagtctac 1800
tttctggtac ctcaaagaaa tcattgttca atcttccata aggaagagat tctttaccag 1860
gctgtgagcc agtgttagat aacttgtgaa tggatataag ttacttttaa caacccctct 1920
tactttttta tttgaatcct ctgaatacct gtcagtattt taaagttggc aatccaggac 1980
attataagta ggatggagca ggagaagaat cctattgaaa ggacaaatta aaatagtaaa 2040
tcctctcctc tcccttctgt aggtactagc tgccttgatt tttttttgtg ggtggggggg 2100
attttttcat tccttattga acttattttg cacaatagtg tttacaaatc ttatacatct 2160
tactttgaca gaagacttat aataccttgt ggtcttgaga ctggttgtct cctttgcctt 2220
tcttcaagga ttttcctttt tctttttacc ccccagtgac cactctgctc tttaggattc 2280
tagtggcagt attcttgtaa cctgttagac gtggatatac ctcttcagga tggcctcctt 2340
gctgtcttta ttgaagagct gcccacccag agtcttgact ctagggcagg ctagtgctat 2400
tgtggtgtca gttgatattt agtttcattt gctgaacccc aaatttgagt gttagttagg 2460
gagcctccat tctgtgggag agtgtggaaa ggcactcttt tttctccata ttagttaaca 2520
agaggtgtct ttaggccctg gatgttacca tctcatttgg tcaagcccct gatacctagt 2580
ttcacatgga tgctaacgtg gatgtaatat agttgtgaca gcctatgcac aaaagcatta 2640
cctgatctta aggtttggta ttctaaccaa agttgacaga ctgtgcgtct gtcttaatgg 2700
atgtcttagg agttggttat tctctctttt tttttgagac ggagtttcgc tgttgttgcc 2760
caggctggag tgcaatggcg tatctccact caccacaacc tccgcctcct gggttcaagg 2820
gattctcctg tcttagcctc ctgagtagct gtgattacag gcatgcgcca ccatgcctgg 2880
ctaatttcat attttttagt agagacagga tttctccatg ttggtcaggc tggtcttgaa 2940
ctcctgacct caggtgatct gcccacctcg gcctcctaaa gtgctgggat tacaggcatg 3000
agccactgca cctggccggt tattctctct ttacagatag ctatagacat cattttagga 3060
8/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
agtgttgcag tctggcattt gtgctattgt tcattctctg tgaaggctgt tcatagttgc 3120
tatagcctgt gtttagtttt gtgatttcat caatcccatc tttctgtgtg agtaatgcat 3180
tctaaacatc ctaccccact ttagaaacgg acgtggggaa cgcttggtca tttaagccaa 3240
caataaattt aggtgaatgt ccctaagtgt ttactgtttt tatccagtca aggatttgct 3300
tttccttgaa catttgtttt aaattctggg gccaaaatgc aaaggagaag ttctattcaa 3360
aggcagtagt tgaaatctat tattttagtt agcctacttg gcatttacta catcggtcac 3420
ttctccaggc tgccctaaat taggttgatg gagtgagaca tgccaaacat ccacctttgg 3480
gaccatagca tagttaaaat taaatgtagt tggaatagct agcattgcag ctacagtagg 3540
gaactgtagt ctagttccct acagaaaacc caaggagtga agggacagga ttttgcctag 3600
gcaaaaatct aagactcgtg ccctcctggt acatggggtt ttaagactga atgtgtaata 3660
ggagcactgc ctttgccaaa tcaaatgagt gacaggttaa ctagaaaatg tgacaatcac 3720
atttcctctt agctcaaata attctgtttt tccaaagctt tagcagctta attaaatctg 3780
ttggactggg ggaggagaga gctgttctct agtggttaac atggtattct ttaagaagaa 3840
aaaacaaagc caaagaaaac tcattatctg gcatgttcgc cttaaagatg gtactgggta 3900
gaatctggag ttttcatctc ttttcaaagc tgcatatctc tcatatttgg tattggcctc 3960
taagtctaat attgcagttg gaattcttgc tgtattattt tttaagcaag tgttaggtgc 4020
atttaactgc tttcttcatc catgacgaca ttcccaccat gggggtcttg acaaagcaga 4080
gtaaaaatat gctgtttaca ttgtttactt acaagtaagg agcctgaaat aacctgtagt 4140
ttcgaatgca ggccctgatt tactggcgtt gtcagtttca attatgaaac tgaagtttgg 4200
tgcctcctct ttatcatgtt ttttcccttg tagcagttgt gtttaatgtc attaaaaaga 4260
aataaaagtt ctttgtcagt gacaaaaaaa aaaaaaaaaa aaaaaaaaat tg 4312
<210> 57
<211> 3860
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
~223> Incyte ID No: 1848891CB1
<400> 57
gggactgaga gccgttccca cgtgagaggc tccgcggccg aattcctcgc gtgcagcagg 60
cgcggaccgc ccggcgtccg gccggactga gagccctggt ccggcgcgcg ccgccggccg 120
ggcgggaggc gggggcgggc tcggctgcgc cccgatgcgg cggcgacctc cgggtctgtg 180
agcccggcgc gcgccgtcgg agcccctcgc gcagccgctg gtagcgtccc cccgggcacc 240
cggcatgcgg gcggccgact cgggctcgtg ggagcgcgtc cgccagctcg cggcgcaggg 300
cgagccggcg ccttcctgcg gggcgggggc cgggcccgcg cggcccccgg gacccgcagc 360
ctgcgagcag tgcgtggacg cggcggggec cggcgatcgg ccccgcgccg gggttccccg 420
ggtccgagcg gatggcgact gcagccagcc cgtgctcctg cgggaagaag tgtcgcggct 480
ccaggaggaa gttcaccttc tccggcagat gaaggagatg ttggcgaagg acctggagga 540
gtcgcagggc ggcaagtcct ctgaggtcct ctcggccacc gagctcaggg tccagctggc 600
ccagaaggag caggagctag ccagagccaa agaagccttg caggccatga aagctgatcg 660
gaagcgctta aagggcgaga agacagacct ggtgagccag atgcagcagc tgtatgccac 720
actggagagc cgcgaggagc agctccgaga cttcatccgc aactatgagc agcaccgcaa 780
ggagagcgag gatgcggtca aagcgctggc caaggagaag gacctgctgg agcgtgagaa 840
gtgggagctg cggcgccaag ccaaggaggc cacagaccac gccacggcac tgcgctccca 900
gctggacctc aaggacaacc ggatgaagga gctggaggcc gagctggcca tggccaaaca 960
gtccttagct acgctgacca aggacgtccc caagcggcat tccctcgcca tgccgggcga 1020
gacggtgctc aatggcaacc aggagtgggt ggtgcaggcg gacctcccgc tgaccgcagc 1080
catccggcag agtcaacaga ctctctacca ctcacacccc cctcaccctg cggaccggca 1140
agcggtcagg gtgagcccct gccactcccg gcagccctct gtcatctccg acgcatctgc 1200
cgccgaaggc gaccggtcgt ccacaccgag cgacatcaac tcccctcgac accggacaca 1260
ctccctctgc aacggcgaca gtcccggccc agttcagaag aacctgcaca accctattgt 1320
acagtcacta gaggatcttg aagaccaaaa acggaaaaag aagaaagaga agatgggatt 1380
cggctccatc tcccgcgtct tcgccagagg gaagcagcgg aagtccctcg accccggcct 1440
ctttgatggt accgcccctg attattacat agaggaggac gcggactggt gatacgcgct 1500
cccctgcgcc tgctgcccgc aggcgtgtct gtgcgtgtgg gcgtgtgtgc aagcgagcgt 1560
gggtgcgcgt gtggccgtgc gtggggtgcg tgtgcacgtg tgcgctggca cacatgggtg 1620
ctgggtgtgg ccgagcgcct ctaacaagtg aaaacacgag tgtgaacctc tctcccctgc 1680
gtcgccacct ctgtaattga tgtacatacc gcaaaccgtg tgtgaacctg tcaactctct 1740
gtcgtctttg gagcgataca gttgtgttgt taatctggtt tattattttt cttcagtgtt 1800
tggtttttct ttttcttttt gtttggttcg tcggtttgtt tttgtttttg tttttttccc 1860
59/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
cctttctcct CCCCtCCtCC tttttatgaa acttgaaaac ttgaaggact gctgtgtatt 1920
tgtaaataac aaaactattg tgcactctgt gcttgtaaat gtccctcgtc caaaccgcta 1980
ctcctggagc ccgtctggca gaggatgtgg tctgtttttg atgtcccccc tcccgccccc 2040
ctggtgtgaa cgtgtgggac cagaccctgt cctgggggtg cgcccaagtc actttaacca 2100
caaaacgcca tcgtcgtcag ggtaagctct gctctctaca aagactcgcg agccgggcca 2160
aggggccttg tcttggctgg gtttgtcaga ggtcaaaccg gctcttttaa acggcctacC 2220
agtttttaaa ttgcattgcc gtttctttct ttatgaaaaa aaagaaaaaa agaaaattgt 2280
ttcatttaat ttatttgcac aaatgctgaa aacttattct atctaaatta ttacataaat 2340
attggaatgt ctatttttcc atggggtggg cgggaggtgg gtgtctctgt tgacttgtct 2400
gttctgttac catgctgcta cccaactgtg caaagtagtt tagggtggcc agaacccagg 2460
gaccattgga tttcaaagct tgctttttct gttggttctt ctctctcctt ctctctctgt 2520
ctctcccatt ctcctgccca tgcatgaaag gatcctccac cttcttccca cccagagctC 2580
cctccaggcc tttctctata tatttattta tctgacatac agaacacgac tttagtgagc 2640
agagtgctga cagtcatggt ccccttcttt gggtctgtct tttgagagag gttgagttca 2700
gggcaagaca gcctgccaca catccaaggg tacgaccagt gggaccctgg cctgagtctg 2760
ttctccgagg ggcctccagc agcttctgtt cctccctgca gctgtgtctt tcttgtcctg 2820
ggtttaggat gcaggtgggc caggcaggtg ttgttagggg aggcacccac ttcaaaagga 2880
gggccacagt ggggacacag agtccagcac ctgagccctc caccctcccc attctggttg 2940
gttccatcag ccacacttag aatctcccag gatcccttgg actccgtccc ccaacttgtc 3000
acagcagcct gagcctaccc accccaggag acaagagctg gcaggaacat ctgcctatat 3060
ggggggtggc gttggcccca aagaggcttc accagcaaag gaactgtgtg tattttaatg 3120
ccaggggacg gaggatgtgt gagctgttca gcaaggtctg cccaaggcct aaaactcaat 3180
ttccttattc ttttgcttct gctcgtcctt aacaactaac agctcaaccc acacctctag 3240
acaacagtag tcgtgctttc tgctaacggt gacattttca gctcttaaaa agaagcaagg 3300
agattttcaa atgctagagt atctctatca gaaggcatag gacattgtgt ccaaagtctc 3360
aagaacaaac aacactttcc ttctgacctg gtccaaaagt cagcaaacag caagcaggca 3420
gaggccctca taaggacttt tctgtcctcc tttgaccaaa ctgtttaacc gagcctaggg 3480
gtgacgggga gcgacccaag ctggcatctt tctctacgga gacagatttt agaaaatact 3540
tttcttgccc atgaatttct tttctggttg atttttatca ttttcccttt acttacaaga 3600
aaataagatt gcaaccactc ctgctaatga tttagtagtt ccttttcatt tcagtttttg 3660
taaattagga gataattcta agagttacta aaggatgatt tatttaagag aactacgtca 3720
aatagcgaat gagttatggg taacattaga cgaaaataac ctttcccgtg ggaaaggttt 3780
ctcgaaggca tggatgcaaa tataaaatat taaaaaaaat ctaaataaag cttattttaa 3840
aatatgaaaa aaaaaaaaaa 3860
<220> 58
<211> 3742
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 2500251CB2
<220>
<221> unsure
<222> 3725
<223> a, t, c, g, or other
<400> 58
cgcacgcata acagccgtgg tggttatggc tggtctgagc ggcgcgcaga tccccgacgg 60
ggagtttacc gcgctagtgt accggctcat ccgcgatgcc cgctacgccg aggcggtgca 120
gctgctgggc cgagaactgc agcggagccc caggagccgt gccggcctgt cgctgctagg 180
ctactgctac taccgcctgc aggagttcgc gctggcggcc gagtgctatg agcagctggg 240
ccagctgcac ccggaactgg agcagtaccg cctgtaccag gcccaggccc tgtacaaggc 300
ctgcctttat ccggaggcca ctcgggtcgc cttccttctc ctggataacc ccgcctacca 360
cagccgggtc ctccgcctgc aagctgccat caagtatagc gagggcgatc tgccagggtc 420
caggagcctg gtggagcagc tgctgagtgg ggaaggggga gaagaaagtg gaggcgacaa 480
tgagaccgat ggccaggtca acctgggttg tttgctctac aaggagggac agtatgaagc 540
tgcatgctcc aagttttctg ccacactgca ggcctcgggc taccagcctg acctttccta 600
caacctggct ttggcctatt acagcagccg acagtatgcc tcagcactga agcatatcgc 660
tgagattatt gagcgtggca tccgccagca tcctgagcta ggtgtgggca tgaccaccga 720
gggctttgat gttcgcagtg ttggcaacac cttagttctc catcagactg ctctggtgga 780
60/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
agccttcaac cttaaggcag ccatagaata ccaactgaga aactatgagg tagctcaaga 840
aaccctcacc gacatgccac ccagggcaga ggaagagttg gaccctgtga ccctgcacaa 900
ccaggcacta atgaacatgg atgccaggcc tacagaaggg tttgaaaagc tacagttttt 960
gctccaacag aatccctttc ctccagagac ttttggcaac ctgttgctgc tctactgtaa 1020
atatgagtat tttgacctgg cagcagatgt cctggcagaa aatgcccatt tgacgtataa 1080
gttcctcaca ccctatctct atgacttctt agatgccctg atcacttgcc agacagctcc 1140
tgaagaggct ttcattaagc ttgatgggct agcagggatg ctgactgagc agcttcggag 1200
actcaccaag caagtacagg aagcaagaca caacagagat gatgaagcta tcaaaaaggc 1260
agtgaatgaa tatgatgaaa ccatggagaa atacattcct gtgttgatgg ctcaggcaaa 1320
aatctactgg aatcttgaaa attatccaat ggtggaaaag atcttccgca aatctgtgga 1380
attctgtaac gaccatgatg tgtggaagtt gaatgtggct catgttctgt tcatgcagga 1440
aaacaaatac aaagaagcca ttggtttcta tgaacccata gtcaagaagc attatgataa 1500
catcctgaat gtcagtgcta ttgtactggc taatctctgt gtttcctata ttatgacaag 1560
tcaaaatgaa gaagcagagg agttgatgag gaagattgaa aaggaggaag agcagctctc 1620
ttatgatgac ccaaatagga aaatgtacca tctctgcatt gtgaatttgg tgataggaac 1680
tctttattgt gccaaaggaa actatgagtt tggtatttct cgagttatca aaagcttgga 1740
gccttataat aaaaagctgg gaacagatac ctggtattat gccaaaagat gcttcctgtc 1800
cttgttagaa aacatgtcaa aacacatgat agtcattcat gacagtgtta ttcaagaatg 1860
tgtccagttt ttaggacact gtgaacttta tggcacaaac atacctgctg ttattgaaca 1920
acccctcgaa gaagaaagaa tgcatgttgg gaagaataca gtcacagatg agtccagaca 1980
attgaaagct ttgatttatg agattatagg atggaataag tagttatgac tgatagtggc 2040
ttttttcaaa atggctttct tacgtaccac actttttttt atctgtattt agccttggca 2100
tctttatatt tgtcttattt tgaatcttat ccactttgta agaacaagtt tatgtttgag 2160
caactttttc atttaatcca gaagggtagg gactatgcag tgtaagctgc atcacttctg 2220
ctttcttcct actagtgaca atcatctggt cttgccctca agcaacaatt gctagagtaa 2280
catctttgta taagcaagta accccagata gagttgacgt ttcagctttg ggctgtcaaa 2340
agggtatgtc atggaccaaa gcactgttag tacgggtatg tttgcatttg gtcactgata 2400
tgtaaatgac tgctagccca cggctggacc acttctcaat cagcaaataa agccatgtct 2460
attttgctat ctcagcatag actatgctgt ctgataaatc taattcttaa ctctatttct 2520
ccagtttttt agtcctttaa ctttctggat tgcaacgaag tctagtttag acctctaagc 2580
ccttttagaa gtacaagtat aatgggaatt tcttttcttg gttcttttca ggttatgagg 2640
tttggtcagt gacaaaattt tttttcataa tttggttgat tggttgcttc ttaagtttta 2700
taataaacgt ttttcttcat gttctatttt tgattttaca taaatgattt tgcctccttg 2760
tggatactga catatattaa gtgtggaagc ttattaatat ttttggtttt ttaaaaactg 2820
aaatttttaa tttttacttt ttaatttttt aggaaaaaat aagcactgaa ctgagaatga 2880
gaagaataaa agtatgagtt ccataccttc taattttagg ctgtcagaaa ttcctttatt 2940
ctttgggatt tcacaatcat ttgaactatc agaagccttt acaattactt ttagctgtaa 3000
catccgattc tgtataagcc acatagaaaa aagttgcctt tcttttttta tgacctggat 3060
atataagcaa atcagctagg aaatatataa ttgtatttta tattaatgtt ttctaggatt 3120
ttggcttaca gtaaatgtta gcccctatgg taagtgattg ttattgttgg atgttatact 3180
gattattaat aagaaatttg gatttttgcc tttttacctg gaatttttgc ttacagccgt 3240
agctatgaat atatataggg tggtccccag tctctgttat ggttgcgcat aaattaataa 3300
ttttataagt atttagaaat ggtataattc tcttaacttc ctctttcagt ttttgtacta 3360
atgtttcgtt tttgttcggg aagaggagat tcgcttttaa gtgcttccaa aaaatgatga 3420
accaccgttc cattcagaaa aaaaggaaca gcacaacgca acacgcaaag gagacaacta 3480
tgcggagcaa cttcttgcaa acaacaatgg atgaggggac gcggggccaa ttaaaacaca 3540
caacaacata tcagcagagt ataagacaaa aaaaaaagga gggcgggaaa gataaacaaa 3600
aaacacgata tatggaccac ggcaaacaca aggaggggcc gaggcacaca aatcagcaaa 3660
ataggtagtt atcgagcgcc acagagggca gtaaaaacga ggggggagac cggcggtcac 3720
tttcntgacc ctcctccaca ga 3742
<210> 59
<211> 2160
<z12> DNA
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 55026561CB1
<400> 59
gcaaaatggg cctctcccca actcccctcc tcatccccca gcacaaggca aagggcaccc 60
cttgctgtga gtttgggggc accggctgtc gggggccgac ccagaacttt gcggggtccc 120
61/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
ctgagggacc cgtggcggga ccggccccgg gttcgtcaaa ttccgccgat gctttcagga 180
ggctgattaa acaaacgaca agattggagt gggaaatgag aggctcttaa aagtggaagc 240
tttttgtttg tccatttaca agaaattgga caatgaatca cttcaggaag atggaggtca 300
tcaacctcac caccctacct atgataccag tggatgagca cctggctgtc tcgcttgtcg 360
,cacggaatac aatggtgaag actgtgagga aggagttaga gaacaatcca ccctcatgcc 420
ttattggctc catgcaccag gtgaaccaaa agattgctga cataaatctg cgtaccgagc 480
cgtcggccaa cagcctggca attgagagat ttgagttgga gaagaaggct ttaagagaga 540
aaactcgcag cagtccagaa gacaaagtca agagacaaag gaaatctcag tattcctgca 600
aaggctccga actcagacat gccagatctt ctgttataaa aaggaaaaca gcagataaaa 660
atctgctggc agagctgtac cagtattcca acttcaacag ctccaagcca aacaagcttc 720
cgaatggcgt ggacttctgt gacatggtgg gcaacgtggt ccgggctgag agagactgcc 780
ttagtggcaa gcatttctgt tcaggtagag aattagagaa gtttctctct tcttcttctc 840
caagagccat ctggctggat agcttttggt ggatatttca tgagaggtac cagccaaaca 900
aggagctcca gaataatctg tttgaccgga tagcccagca ctatgcctta cttttgtttc 960
gtgtacccaa gtcccactct gaagaggcgc tcttaaaaag gctgccatca cttctcagca 1020
aagccgtgta caccagcttc tgttgctgct ttccacagtc ctggttcgac acgcacgaat 1080
tcaagtctga catctgtaac acaatgagcc tgtggatttc aggcacctat CCtagCCCdC 1140
agagctatga cagctgggac tactcggaac tagacccaga gcgattccgc agagaagaat 1200
taatgttgta cagaagaaga ctgacaaagg ggagagagtt ttctttgttt gctggtaaga 1260
gagccttctc ccagaagcca gcccagagca ggaaattcta ccaccctcag tcttctagtg 1320
caaattcacc cagtgaaaaa acctcttcgg ccaagcagaa ctcagaaaaa agcttacgaa 1380
tgcagaatac tgcaaaagag catcattgtc agaccctggt cttgaagaaa cctacgcaag 1440
aagtcaagag gatatcagaa gcaagagaat gtgagaatat gtttcctaaa aagtcgtgtg 1500
ctgcctgcaa aagccctgag ctgacttcaa acctcttcaa catttatggg aagagccctc 1560
tgattgtgta ctttctccag aactatgcca gtctgcagca gcacggcaag aatgtgttga 1620
tagtcagaag ggaaaagacc acgagcaccc ctgactgcac cccaacgtat actgatgtca 1680
tcagcgagac cctgtgcagc atgaagaagc ggaaagacaa cctcaatcag ttgtaccagc 1740
atcattggac tgaatggaat tattttgaca agcatctaaa ggagctgcaa gacaacttct 1800
ccagggaaat gaagaatatt ggtccaaaag cagcagatac aaaaaaggca aaccacatgt 1860
tcatcccacc ttcagccgtc aatgaggaat cacctgacaa gaaaactaag ggaagtctcc 1920
aaagagaaat tgagtttaag ggttgttcga acaaaaacca tgggaaaggt agaagtgtga 1980
acatgagggg aaaggaggag agggaaagag aagagaaaca gaagttgaac atttctttcc 2040
actcacttcc aagccctgag gagctccaca acctagaacc aggaagcgcc tacagaatcc 2100
gtgatatttc tgctacgagg tggagtggaa caagaaaata agaaaattgt ggccaaagac 2160
<210> 60
<211> 1840
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7502593CB1
<400> 60
gtcttacaac aaagccaagg aatctcgctg ctgagggcag ttctgtgctt tattatgaag 60
aataatggac gatgatgatg caaagctcaa agcagaaata gaagctgaat tggataaact 120
cagcatttcc tccttggaaa aagaagacat tgagagtgat gcaaaatcag aaacccagag 180
tgatgatagt gatacagatt cagttgaatt accagaatca gttcttcact gtattaacat 240
cataaagaac aggagtaaag ctgttgaaga gctcattctt caggacctgg aagatactga 300
tattttaagc tgtagttatg gagcagtttc taataatcat atgcatttaa gaacaggact 360
atcaactgaa tatgaagaaa gttcagagca attaattaag atattatctg aaatagaaaa 420
agaagaattt atgagaagta aaaccgattg tgccactcct gattttgttc ctgagcctag 480
tcctcatgac ttgcctatgg atgaacatgt tttaccagat gatgctgata taaattttgg 540
atactgtgaa gtggaagaaa aatgtagaca gtcttttgag gcttggcaag agaaacagaa 600
ggaattagaa gataaagaga aacaaactct caaagctcag agggatagag aagaaaaaca 660
atttcaagaa aaaaaaaaaa agcgacattg ctggatgaaa caatttaaag ttgaaaagaa 720
gaaattagag aacattcaga aggtattttg cttttgtttt tcatgtattt ttaaaatcag 780
tagctacctc tgaagaatat tgatatttgg attgtgttgc agattgagct taatagtatg 840
acttttaatt actctCtgat agaggaagaa attgactgtg aattaacagt atttaatatg 900
aggtccttga ttagttttta ttgaaattca ttggaaggtc agacacactt ctgatcttgg 960
gggccagtgg agtctatgaa tttcaagggt catcatggat agcctgaaat ttacaatata 1020
aattgttgtt tttctttata tgcttctaaa cattggaaaa agcaatgaaa aattcttgag 1080
62190
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
tatgcattaa ccttgctctg tggtctggca tatgacatgt aacacatatt ggaaaagtag 1140
ttcaaccatt tagagtcagt ctgcgcccca ccttcatgaa ttaagctact tcctttgaac 1200
tggcaacttt ttggtgtgaa gttttgtact gttttcattt ctgtcgtatg tgttgttggt 1260
gttgttttcc ccctaggaga tccacacaat cagatcatag tggtggtata tcggtattga 1320
gaaactggtg gacttttagg acaccacttg caggtcatga gaaccttaga atatttggat 1380
aacaggcatg actgacttct ctatctttca tgattaaagt tgatttatac attatccaga 1440
acttgctggg tttgagtgtt gcatataaag aacattagct tgttcaacat tttttaatcc 1500
tccatgttga ggacatttca tttcttgact gtgtagaaaa taaacacatt aaaggctggg 1560
cgcggtggct cacgcctgta atcccagcac cttgggaggc cgagacgggt gtataacgag 1620
gtcaggagat cgagatcatc ctggctaaca cggtgaaacc ccgtctctac taaaaataga 1680
gaaaattagc caggcatagt ggcgggcgcc tctagtccca gctacccggg aggctgaggc 1740
aggagaatgg cgtgaacccg ggaggcagac cttgcagtga gccaagatcg tgccactgca 1800
ctccagcctg ggcgacagag caagactccg tctcaaaaaa 1840
<210> 61
<211> 1808
<212> DNA
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7503957CB1
<400> 61
tcttatgcca tgcctctagt gtgccacatt tatgttcacg atctcattta attcttgcca 60
caaccctatc aggaaggtgg gagtcaatca ttttgacaag tctcctgaaa ggaacagcta 120
gcaggaactg aaaccttttt ccatttggtc tcgtggcaaa ggcagagatt gctccagcag 180
ctccacacaa aatgatgtgc tcacgggtgc cctctgaaca gtcttctggt acctctctct 240
tgcctaaaga cggtgcccca ttttcttggg attccttgga tgaggatgga ttggatgact 300
ccttgctgga gctgtcagag ggagaagaag atgatggtga tgtaaattac acagaggaag 360
agattgatgc actgttgaag gaagatgacc catcatatga gcagtcttct ggggaagatg 420
atggtgggca tgttgagaag ggagaaagag ggagtcaaat tctacttgat actccccgag 480
agaaaaattc atcgtacagc ctgggaccag tagctgagac tcctgacctc ttcaaactac 540
ctcagctaag tacatcaagt ggtcatggac cagctcatac taaaccatta aacagacgct 600
ctgtactaga aaagaatctt ataaaagtaa ctgttgcacc atttaatcca acagtttgtg 660
atgctctgct tgataaggac gagactgatt cgtccaaaga tactgaaaaa ctctcttccc 720
ttggagaaga gatgagagaa gatggtctta gcccaaatga aagcaaactt tgtactgaat 780
ctgaagggat cagccccaat aactctgcct ggaatgggcc ccagctctct tcttcaaaca 840
ataactttca acagactgtc tctgataaaa atatgcctga cagtgagaac cctacgtctg 900
tattctctcg gatctcagac cattcagaga ctcctaatat ggagttatcc tgcagaaatg 960
gtggttcaca caagtcaagt tgtgaaatga gatctctggt tgtttccacc tcatcaaaca 1020
aacaggatgt tcttaacaag gattctggga agatgaaagg ccatgagaga agactaggca 1080
aagtcattcc tgttctacaa actaagacca ggactaatgt tccgacgttt tcacagtcaa 1140
atctagaaca gcagaagcag ctttatctca ggagtgtcat tgctcatata gaagacccag 1200
aggacactaa ccaaggtatc tcgggggagc tttgtgcctt gatggatcaa gttcatcata 1260
tgcagcactc aaaatggcag catccttcgg acctcaccac gcgaaactac gcccgccgac 1320
agaaacatct gcaaagatac agtctgactc agtgggttga caggaacatg cgaagccacc 1380
atcggttcca gcgtctccca gacttctcgt acagttaatt tgtgtcatcc catcagcaat 1440
gaaggtccct atccagggtc ctgcttggag cagcatttca tgttcttttg ctgttttgtg 1500
ctttgccgat tttggatttt atttttcaca aaatttttat ttaaaaaact cgtcaccttt 1560
tggaaatgcc cattgccgac ttgaattttt tggtatgaag tccctcctga ttttgtgtgt 1620
gtgtgtctgt gtttaagcaa gcgttcggtt ggtatagtta cggtttttaa tttaaattga 1680
aggtagctgc tcctgaaagc cagcattaag gccagaacac ccaggttcaa gcaaaagacc 1740
cacctctctg tcagaggcaa agtctaattt ctggtactca aagaaattca ttgtccatct 1800
tccataag 1808
<210> 62
<211> 3941
<212> DNA
<213> Homo sapiens
<220>
<221> misc_feature
63/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<223> Incyte ID No: 7504415CB1
<400> 62
ggcggcggtt ttggctgtgt gaggaagacg gaagagacgg cggcggaggg aaaccgactt 60
ccactagtcc gggtcgcttg ggcggccggg ggccctcaga gtctcccggg cagtggtagc 120
agttgcagca ggatcaggcg cctgtcggct tctgacgttt aaaacagggg gagcggaagg 180
gagccactgg ccgcggtggc agggccaggt ataaggaagg aaaatatggc ggcggcggcg 240
gcggcctgag gaggcggcgg cggcgcggga agctgcttcg cggagatcat ggcggaggcg 300
ggagcagggc agtgacggga gccccgagtt cctagcgctg cggggcggga ggctacgaag 360
cgctgcgcgg ccccctcggg gctgccgggc gccgggctcg ccaggcctgg acaatagcgc 420
cggggagccg gaggcgagga aaggcggcgg cccagagctc ggtccctgga gcgggccatg 480
caggcggcgg cgcggccccg cggcgcCCag cggcggcagt gaggccgggg agCCCtCCgC 540
tcgcgggcgc cctcacgcct cgcccctcgc ctctccaggg cccctttcct gggcgtctac 600
tggcggggcc cccgccccgg ttcccgggcg gcacgatgac cgacacccgg cggcgggtga 660
aggtgtacac gctcaacgag gaccggca°gt gggacgaccg gggcaccggg catgtgtcgt 720
ctggctacgt ggagcggctg aagggcatgt ccctgcttgt cagggctgag agcgacggtt 780
ctctactttt agagtcgaaa ataaatccta acactgcata ccagaaacaa caggacactc 840
tgattgtgtg gtctgaagca gaaaattatg acttggccct tagctttcaa gaaaaagctg 900
gatgtgatga aatttgggag aaaatatgtc aggttcaagg aaaggaccct tccgtggaca 960
tcactcagga ccttgtggat gaatctgaag aggagcgttt tgatgatatg tcatcgccag 1020
gcttagaatt gccatcttgt gaattaagtc gccttgaaga aattgcagaa cttgtggcat 1080
catCtttacc ttcacctctt cgtcgtgaaa aacttgcact ggcactagaa aatgagggtt 1140
atattaaaaa gctcctggag ctttttcatg tgtgtgaaga tttggaaaat attgaaggac 1200
tgcaccactt gtatgaaatt atcaaaggca tctttctctt gaatcgaact gctctttttg 1260
aagttatgtt ctctgaagaa tgtataatgg acgtcattgg atgtttagaa tatgatcctg 1320
ctttatcaca accacgaaaa cacagggaat ttctaacaaa aacagccaag tttaaagaag 1380
tgattcccat atcagatcct gagctgaaac aaaaaattca tcagacatac agagttcagt 1440
atatacaaga tatggttcta ccaactcctt cggtctttga agaaaacatg ttatcaacac 1500
ttcactcttt tatctttttc aataaggtag agattgttgg catgttgcag gaagatgaaa 1560
aatttctgac.agatttgttt gcacaactaa cagatgaagc aacagatgag gaaaaaagac 1620
aggaattggt taacttttta aaagaatttt gtgcgttttc ccaaacgcta cagcctcaaa 1680
acagagatgc ttttttcaag actttgtcaa acatgggcat attaccagct ttagaagtca 1740
tccttggcat ggatgataca caggtgcgaa gtgctgctac tgatatattc tcatacttgg 1800
ttgaatataa tccatccatg gtacgagagt ttgtcatgca ggaggcacaa cagaatgatg 1860
atgatatttt gctcatcaac ctcattatag aacatatgat ttgtgataca gatcctgaac 1920
ttggaggagc agtccagctt atgggcctgc ttcgaacttt agttgaccca gagaacatgc 1980
tagccactgc caataaaaca gaaaagactg aatttctggg tttcttctac aagcactgta 2040
tgcatgttct cactgctcct ttactagcaa atacaacaga agacaaacct agtaaagatg 2100
attttcagac tgcccaacta ttggcacttg tattggaatt gttaacattt tgtgtggagc 2160
accataccta ccacataaag aactacatta ttaataagga tatcctccgg agagtgctag 2220
ttcttatggc ctcgaagcat gctttcttgg cattatgtgo ccttcgtttt aaaagaaaga 2280
ttattggatt aaaagatgag ttttacaacc gctacataat gaaaagtttt ttgtttgaac 2340
cagtagtgaa agcatttctc aacaatggat cccgctacaa tctgatgaac tctgccataa 2400
tagagatgtt tgaatttatt agagtggaag atataaaatc attaactgct catgtaattg 2460
aaaattactg gaaagcactg gaagatgtag attatgtaca gacatttaaa ggattaaaac 2520
tgagatttga acaacaaaga gaaaggcaag ataatcccaa acttgacagt atgcgttcca 2580
ttttgaggaa tcacagatat cgaagagatg ccagaacact agaagatgaa gaagagatgt 2640
ggtttaacac agatgaagat gacatggaag atggagaagc tgtagtgtct ccatctgaca 2700
aaactaaaaa tgatgatgat attatggatc caataagtaa attcatggaa aggaagaaat 2760
taaaagaaag tgaggaaaag gaagtgcttc tgaaaacaaa cctttctgga cggcagagcc 2820
caagtttcaa gctttccctg tccagtggaa cgaagactaa cctcaccagc cagtcatcta 2880
caacaaatct gcctggttct ccgggatcac ctggatcccc aggatctcca ggctctcctg 2940
gatccgtacc taaaaataca tctcagacgg cagctattac tacaaaggga ggcctcgtgg 3000
gtctggtaga ttatcctgat gatgatgaag atgatgatga ggatgaagat aaggaagata 3060
cgttaccatt gtcaaagaaa gcaaaatttg attcataata atggcaacgg cctaggatca 3120
gtacctgttg aaaaaaactg gttctccacc cctcccccat acaaaatcca caaaaaagcg 3180
cagtggtctc ttgtgaatga ctgacacaga tcagcctctt acacttgact tctgctcatc 3240
aagtgccaat tcaatggagc aggaggaggg gatatcatat atttagggga aagacttaag 3300
cctttgagct ctccagcttg gaccacacat tgcccttttc tcagggaagg aaatggaaac 3360
aaaaagccaa cagggcaggg gttttgtaag tggaactctg gattgactgg tcagttgcta 3420
caatcagaat atgctttctt ggaccatgtt tgagactcag aagaatgggc ctttctgcca 3480
taattcttca ctagtcaaga atgccagcag tttctttgta taaagagacc tgcctttaaa 3540
atcatacatt ctgaacattt tagtcaagct acaacaggtt tggaaaacct ctgtggggga 3600
64/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
ggggcgagta taaagttttc ctctttttta actgttccct ttgcccttca aactgcagat 3660
attttttttt ttaagtgggg acttctccct acttgattaa agattgagtg gaattctaga 3720
tgtggtcatt tgtgtcataa ttttttggtt ttatttggtt ttggattttt ttttccctcc 3780
cctgagttgt ttgcttagtt gtggcagttc ttcttttttg gcggaccttt aaaacttttt 3840
tggatgtatt tataccctaa cgttgtgctg gtccctgttt tcccatgtgt attttggtct 3900
acatccagtt cttattggtt cagagtttct gacgctgctc a 3941
<210> 63
<211> 1933
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7504074CB1
<400> 63
cctaaggtag cgacttgctt tctgacgagc cacacgtttg cttcttccct gtgttcccag 60
ctggagggac atgagtgtcc ctgggccgtc gtctccggac ggggccctga cacggccacc 120
ctactgcCtg gaggccgggg agccgacgcc tgacagaacc aagttgtatt gttagacaca 180
ctggaacaag agatttcaaa atttaaagaa tgtcattcta tgttggatat taatgctttg 240
tttgctgagg ctaaacacta tcatgccaag ttggtgaata taagaaaaga gatgctgatg 300
cttcatgaaa aaacatcaaa gttaaaaaaa agagcactta aactgcagca gaagaggcaa 360
aaagaagagt tggaaaggga gcagcaacga gagaaggagt ttgaaagaga aaagcagtta 420
actgccagac cagCCaaaag gatgtgaaaa gttgtgtttg tgtgttttct tctcctgtcc 480
catatttggg ttatgatgac tcaagtgtag actgaagttg aggtagtgcc ttatgccatt 540
atgtcatatg ttgaaatcct tattccggta ttactgtgtc tccatgcctt ttttccaagt 600
agcagacgtc atgttgcatg gtttttgata tttatatgta agtttttcaa attttgctta 660
attttaaaat ttattatttt gatcttgaat tatttataaa ctggaaagtg gtttgattat 720
tgtgagtcaa aactctaagt ggttaaaaat tagtatgaat tttttagctt cttaatgaat 780
atggatttaa aactctccag ttcttatttt atgaaatgac ttgcctttct ggtaatacaa 840
tgctgatttt ttagtaattg ccttttcatt actttgttaa gaagaaatgc cagctgttta 900
atcacaccta cccctggaaa agaggtaaac cttttgaaca gttgaatttc atcagaagct 960
ctatagcttt ttggtgagag gaagtgatac tctttattac aagaaacaag gaattaacaa 1020
aaataaggaa cttgttgcta cctttctttt ctgttgaaca ttaaaaataa tttggtttgt 1080
ggactgggca tggtggctta tggaaaagag gtgagccttt gtgaagaaca taatggaaaa 1140
gtgcatgtac gggaaataac agcactttag gtgtagttca ggtgagcaag atgccagata 1200
gtttctacta accatatact Ctttctgctt cttagcgtcg ttgtcactgc ttctgtggct 1260
tttgggttca aagtagatta tatatattcc taaagcttgg tggaggtagt ggagctatca 1320
cacactcaaa tgagcttgta tatttataat acaatatagt attatagtaa cagttatcat 1380
acactgagat atagtttctg tgcttgtaaa aaattcttga agtgaggccg ggtgcggtgg 1440
ctcatgcctg taatcccaac gctttgggac gccgaggcgg gtggatcatg aggtcaaaag 1500
attgagacca tcctggccaa catggtgaaa ccccgtctct actagaaata caaaaattag 1560
ctgggtgtgg gggcgtgcgc ctgtagtccc agctacttgg gaggctgagg caggagaatt 1620
gcttgaaccc aggaggcaga ggttgcagta agccaagatc gtgccactgc actccagcct 1680
tggtgacaga gcaagactcc gtctccaaaa aaaaaaaaaa aaaatttctc tgaagtggaa 1740
ctgatgtaca ccaaataccc actttcatag cataattaca gggatggtga ccaattcagt 1800
acaatagctt caccaaaaga ggcagcgtgg taccttttag gtgtttctag aatctcttct 1860
ggatattcta tgtaagaaat tcatcgttcg agggaactac gagggtttca ttgtctaaaa 1920
catcacgcaa gcg 2933
<210> 64
<211> 4460
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7502257CB1
<400> 64
gcggcgcgaa cggcagctag gagggttgct ccgggettgg tgctcactgc gacttcccgc 60
gcagggcccg gtcggactag gacccgcggc ctgagagacg ctggaggatg cggacgcgga 120
65/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
ggccgcctgg ggtagcggcg gcgggagtcc tggcgctctg caggtcagaa gttgagtagc 180
aggggcctag gagggctcga agccttcaca gcgatggcag agaagcgacc cctgagaacc 240
ctggggcctg tgatgtatgg caagctgccc cgcttagaga cagactccgg gctcgagcac 300
agcctgcccc actctgttgg taaccaggat ccctgcacct acaaggggtc ctacttctcc 360
tgccccatgg cgggtactcc taaggccgag tctgagcagt tggcgtcctg gaccccatac 420
ccacccttgt actctaccgg tatggcagga cccccacttc aggcagacaa cctgctgacc 480
aactgcctgt tctaccgctc gccagcagaa ggccctgaga agatgcagga ctccagccct 540
gttgagctcc tgcccttcag tccccaggct cactcctacc caggcccacc actggcagca 600
cccaaacctg tctaccgcaa ccctctgtgc tatgggctct caacttgtct gggggaagga 660
gcagtgaaga ggccactgga tgttgactgg actctggcga ctgggcccct gttgccctca 720
gctgacccac cctgctctct ggccccagct cctagcaagg gccagactct ggatggcacc 780
ttcttgcggg gggtgccagc tgaggggtcc agtaaagact cctcagggag cttctcccca 840
tgccagccct tcctggagaa atatcagacc atccacagca cgggcttcct ggcctccagg 900
tacacaggtc cttaccctag gaactccaag caagcaatgt ctgaggggcc Ctcaagtcct 960
tggacccagc tggcccagcc cctggggcca ccctgtcagg acaccgggcc cacccactac 1020
ccaccacccc accacccacc accccaccct ccacaggccc tgccttgccc tccagcctgt 1080
cgccacccag agaagcaggg cagctacagc ccagcactcc cactgcagcc tctggggggc 1140
cacaagggga ccgggtacca ggctggtggg ctgggcagcc cctacctgag gcagcaggca 1200
gcccaggcac cttacattcc cccactgggg ctggacgctt acccctaccc ctctgcccct 1260
ctcccagcac cctctccagg cctcaagctg gagccgcctc tcactccacg gtgcccattg 1320
gactttgccc cccagacact gagttttcct tatgcccggg atgacctctc tctctatgga 1380
gcatcccctg ggcttggagg gacaccacct tcccagaaca atgtgagggc tgtgccacag 1440
cccggtgcct tccagagggc atgccagcct ttgccagcga gccagccctg ctcagagcct 1500
gtgaggcctg cacaggaagc cgaagagaag acctggctgc ccagctgcag gaaagagaag 1560
ctccagcccc ggctcagtga gcactctggg ccgcccatcg tcatccgaga cagtccagtt 1620
ccctgtaccc ccccagcact gcccccctgt gcccgggagt gccagtctct tccacagaag 1680
gaggacgcaa ggccacccag ctctccacca atgcctgtca ttgacaatgt cttcagcctg 1740
gccccctacc gtgactatct ggatgtgccg gcacccgagg ccacaactga gcctgactct 1800
gccacagctg agcctgactc agccccagcc accagtgaag gtcaggacaa aggctgcagg 1860
gggaccctgc ctgcccagga gggcccctca gggagtaaac ccctaagggg ctcacttaag 1920
gaggaggtag ccctggattt gagtgtgagg aagcccacag cagaggcctc ccctgtcaag 1980
gcttcccgtt ctgtggagca tgccaagcct actgcagcca tggatgtgcc agatgtgggc 2040
aacatggtgt cagatctgcc aggcctgaaa aagatagaca cagaagcacc aggcttgcct 2100
ggggtgccag tgaccacaga tgccatgcca aggaccaact tccacagctc tgtggccttc 2160
atgttccgaa agttcaagat cctccgtccg gcacctttgc ctgcagccgt ggtcccgtcc 2220
acgcccacct cagctcctgc tcccacacag cctgcaccca cccccacatc tgggcccatt 2280
ggactgcgga ttctcgctca acagcccttg tctgtgacct gcttcagcct ggcactgccc 2340
agccctccag ccgtagctgt~ggcctcccct gcccctgctc cagctccatc ccctgctccg 2400
gctcgagctc aggctccagc ttcagcccgg gatccagctc cagctccagc tccagttgca 2460
ggccctgctc cagcatctac ttcagcccca ggggactccc tggagcagca ttttacagga 2520
ctacatgcgt ccctgtgtga tgctatttct ggctccgtcg cccactctcc tccagagaag 2580
cttcgcgagt ggctagagac ggctgggccc tggggccagg ctgcgtggca ggactgccag 2640
ggtgtgcagg ggctgctggc caagctgctg tctcagctgc agcgcttcga tcgcacccac 2700
cggtgcccct tcccccatgt ggtgcgagct ggcgccatct tcgtgcccat tcacctggtg 2760
aaggagcggc tcttccctcg gctgccaccc gcttctgtgg accatgtgct gcaggagcat 2820
cgtgtggagc tgcggcccac cacgctgtcg gaggagcggg cactgcggga gctcgccctg 2880
ccaggctgca cctcacgcat gctgaagtta ctggcgctgc gccagctgcc ggacatttac 2940
cccgaccttc tcggcctgca gtggcgcgac tgtgtacgcc gccagctggg tgagcatggg 3000
gcagccccag tggccaccgg agctgtgtga gcaagtgaca ggtgactttg acactgaggc 3060
tggagctgtg tcctcctcag agcccactgt ggccagagat gagccagaga gcctagccct 3120
ggctcagaag tcaccggccc ccaaggtcag gaagccaggc aggaagccac caacccctgg 3180
cccggagaaa gcagaggcag ctgctgggga agagtcctgt ggtgcctccc ctacccctgc 3240
taccagtgcc agcccacctg gccccacact gaaggcccgc ttccgcagtc tgctggagac 3300
cgcctggctc aatggcctgg ctctgcccac ctggggccac aagtcctaaa gaccagacca 3360
gCCCtCdCCC tgcccacagc tgctggacag ccagagccat cacctgtagc actggttgcc 3420
agtgctgtgt gtatagcagt CdCtCtCCa.C CCttCCCttC tgcctgccca gctgccccgg 3480
ggccacgagt ggatgctggg gctgtggctg ctcccctgga ggggttccat ctctgaccct 3540
gtggcccatt cagggtgggc tgaagagccc ctgagctttt aacgtgaggg tctttattgg 3600
ataggactac tccctatttc ttgcctagag aacacacatg ggctttggag cccgacagac 3660
ctgggcttga atcccggctc gtgttcttgc tgcaggacct gggcaagaaa cttcacctct 3720
gctgagccct cattccccat gtgtaaaatg ggacaacgca acctacctca cagggttgtt 3780
gtggggatgc tgcctgatac ataccctgtc accatttggt ctctgcttcc tctctgggac 3840
agggcctaga attggaggca gagaaccttc ctatagaaag tcttcgtgtg tcctaggact 3900
66/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
tggctatcgt agagtggtac cttaggcagt ggatgtgact cacactttca ggagtcaccc 3960
cccagcattt ggggttgggt tggccctact ccagcctgga gctccctgag ggagcctgca 4020
ctccctgctc ccaatccccg ctactggtgc agggatgcag cctggagctg gcgtccttgt 4080
tctgggcctg ctgctgccgc caccccagga ggccccaggc ctgtcctgaa ttgacatcag 4140
tgcttccctg aactgcctcc cccacccctg gcattatccc aggaaactta tgttttctag 4200
aagctaagca gctgctggga ctcagggact ggtgcaggta ggctgagtgg cagctcagtc 4260
ctagaaggtc tctgaagatc tggactgagg accctgctac tccccaagcc agagcccatc 4320
agccaggcct gctgtgagcc acctgcctgt ggagtgctga gctcaaccaa aggctggcaa 4380
gctctgggcc tcatttaagg gattctgatg agccgatggg ccctggaggc agcccattaa 4440
agcatctggc tcgtttctgg 4460
<210> 65
<211> 1214
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 1315136CB1
<400> 65
gaggaggagt ggccctgatg aggacccgta gggcttcaac cgacccccag ctcaaaaaca 60
cttcggcttg ggggcggtgc caaggctgtg agtgcttcca acacttcggc ttgggggcgg 120
tgccaaggct gtgagtgcgg aagcctgccg gaaacctggg gacggagcga gggaaatgac 180
gcctgggagc gacaacaggg tgggtggggc tgctgactcc gcccccgaag gaaagggtta 240
acggtttccg gtagcggcgt ctaggagggg cgggggaaag gaggcggcag ccaggctgtg 300
tcccctgacc gttggagcgt CtgCgaCCCC CgCatCCCCg caccctcaag gcacctccaa 360
agatgatgat gggttgtggg gagtcagagc tgaagtcggc ggacggggaa gaagccgcgg 420
cggtcccggg gccacccccg gagccccaag tcccgcaact ccgagcccca gtgcccgagc 480
ccggcctgga cttgagcctg agcccgcggc ccgacagccc tcagccgcgg cacggcagcc 540
ccgggcggcg gaaggggcgg gcggagcggc ggggcgcggc tcggcagcgg cggcaggtcc 600
gcttccgcct gacgccgccc tccccggtgc ggtccgagcc gcagcctgcg gtgccgcagg 660
agctggagat gcccgtgctg aagagcagcc tggccttggg cctggagctg cgggccgcag 720
ccgggagcca ctttgatgct gcgaaggccg tggaggaaca gctgagaaag tcgttccaga 780
tccgctgcgg cctggaggag agcgtgtccg aggggctgaa cgtgccgcgc tccaagcggc 840
tcttccggga cctggtgagc ctgcaggtgc cggaggaaca ggttctgaat gccgcgctca 900
gggagaaatt ggctctcctg ccgccacagg ctcgagcccc gcacccaaag gagccacctg 960
ggcctgggcc agacatgacc atcttgtgtg acccagaaac gctattttat gaatctccac 1020
acctgaccct ggacggtctg ccccctctcc gacttcaact ccggccccgc ccttcagagg 1080
acaccttcct catgcaccgg acactgaggc gatgggaagc gtagacccca aagatccctg 1140
gagggctagt tcgtattttt gtgttaaact atttgttaga ataaagtaat tttgctaata 1200
aaaaaaaaaa aaaa 1214
<210> 66
<211> 2843
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 1379785CB1
<400> 66
agcgaccggc acagcctgcc ccgtcccgcc tggcacgccc tgattagcgc cgggcacgac 60
acgcaccgcc ccgcccggca ctccctgctt cgcccacgac acgcccctcc ccgcccggca 120
aactgcccgg agccaaggcc ccgcccccag agagaacctg ccccgcctcc ctggcccggg 180
ccccgcctcc ttggcaagcg agtggcgctt tcaccttagc aaccagcgcg gctcccacca 240
tggctgaaga agaggaaact gctgctctca cggagaaggt tatccggacc, cagagggtgt 300
ttataaacct gttggattcc tacagcagcg gaaacatcgg gaagtttcta tctaactgtg 360
tagttggggc ttcgcttgaa gaaattacag aggaagagga agaggaagat gaaaataagt 420
cagctatgct ggaagcttcc tcaaccaaag cgaaggaagg cacattccag attgtgggca 480
cgctgtccaa gcctgacagc ccgcggcctg actttgcggt ggagacgtac tctgccatct 540
ctcgagaaga ccttctcatg cgcctgctgg agtgtgatgt tattatttat aacatcactg 600
67/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
agagctcaca gcaaatggag gaagccatct gggcagtctc tgcactcagt gaagaagtca 660
gccactttga aaagcgaaag ctatttattt tactgtcgac ggtgatgact tgggcgcgct 720
ccaaagccct ggaccccgag gattctgagg ttccattcac tgaagaagat tatcgaagaa 780
gaaagtctca tcctaatttt ctggaccaca taaatgctga aaaaatggtt ctcaaatttg 840
gaaaaaaggc cagaaaattt gcagcatacg tagttgctgc tggactccag tatggagcgg 900
aaggaggcat gttacacaca ttttttaaga tggcttggtt gggcgagatt cctgcattac 960
cagtttttgg cgatggaaca aatgtaattc caacaatcca tgttcttgat ctagcaggag 1020
tgatacaaaa cgtcatagat cacgtgccaa agcctcacta cctggttgct gtggatgagt 1080
ctgttcatac cctggaagac atagtcaagt gtatcagtaa aaatactggc cctgggaaaa 1140
tccagaaaat acccagagaa aatgcatacc taaccaagga cttaacgcaa gattgtcttg 1200
accatttact ggtcaactta agaatggaag cgctctttgt gaaggagaat tttaatattc 1260
gatgggctgc ccaaacagga tttgtggaaa atatcaacac tatcctcaag gagtacaagc 1320
aaagcagagg attgatgcca atcaagatct gcattcttgg tccccctgct gtgggaaaat 1380
ccagtattgc taaagaattg gccaactact acaaactgca tcacatccaa ctgaaggatg 1440
tcatttctga agccatagca aaactggagg cgattgttgc ccctaacgat gtaggggaag 1500
gagaagaaga agtcgaagag gaagaggagg aggagaatgt ggaagatgca caggagctcc 1560
tagatggcat caaggagagc atggagcaga atgcaggtca actagacgat caatatataa 1620
ttagatttat gaaagaaaag ctaaaatcaa tgccttgcag gaatcaaggt tatattttgg 1680
atggattccc aaagacctat gatcaagcaa aagacctgtt caatcaggaa gatgaggagg 1740
aggaagatga tgtcagaggc agaatgtttc cctttgataa attaattata cctgaattcg 1800
tttgtgcact ggatgcttcg gatgagtttc tgaaggagcg tgtgataaac cttcctgaga 1860
gcatcgtggc ggggacccac tacagccaag accgattcct ccgggctctg agcaactacc 1920
gggacatcaa tatcgacgat gagactgtct tcaactattt tgatgaactt gaaattcacc 1980
cgatacatat tgatgtagga aaacttgaag atgctcagaa tagacttgct atcaaacagc 2040
tcatcaaaga gattggggag cctcgaaatt atggtttaac agacgaagaa aaggcagaag 2100
aggagcggaa ggctgcggag gagcggctgg ccagggaggc tgctgaggaa gcagaacgcg 2160
agcaccagga ggccgtggag atggcagaga agatagctcg ctgggaggag tggaataaac 2220
gactggagga agtgaaaaga gaagaaagag aattactgga ggctcagtca attcccctga 2280
gaaactattt aatgacctat gtgatgccaa ctcttattca gggcctgaat gaatgttgca 2340
acgtccgacc cgaagaccct gttgattttc tggcagaata tctcttcaag aacaatcctg 2400
aagcacagtg aaacttgaaa gatctggtat tatctacctt tacagaacca cagatcactt 2460
attatacttt gaaaaattgc tttgaaaaat gcttttccag ttcttagaaa attctttttt 2520
tgtagacaaa tattctataa actagaatct ctattaaaag ctatatgaca tgactatgtc 2580
attaaaacta tatttgaaat gtaaattgat aaagacattt gtgcatagct catgagacaa 2640
atactgattt aatattttat tctttagtca gatctaaata taccgcttct gtacactaat 2700
gtttataggt atttatagca tgaagaaaat cagactatat attgtagact atgtattatt 2760
ctaacatgta ggctaattta catgacttgt tatcgcccca ataacaatgt tattagaaat 2820
ggaaataaat tgaagtgatt tat 2843
<210> 67
<211> 1021
<212> DNA
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 2011166CB1
<220>
<221> unsure
<222> 8
<223> a, t, c, g, or other
<400> 67
agcggagntg ggagcctgat ggaggacaag tagggcctcg aggacaggtg cgtgacagaa 60
gcacaggaaa aaaaagaaaa atgaagaaat aaaaacacga gttcatcagt aaagaggtac 120
cctggcagca taaatattat gataagctaa aagctggaat catctggaaa aataaataag 180
actcctcatg tccttttcgg tccataacca gaagggcagc aaaaggcctt tgccactgga 240
acctcttctt tttctccaag tcccacgtag caattacctg cactttcaag aagagaaaca 300
acgactacac ctaaagaaat tccttcttga taggatgttt ctagtggcca agatacaagc 360
aaatgtagaa agaaaagatg ttgctgacta ctatgaacaa atgtttcagt cagttttgaa 420
acatcaccta ggagaagcag tgacaggatt gctgctcatc tatcccactt ccattctgca 480
tatcctcgag tcctccagcg acactctcta caaagttctt ttagattata ttggccatgt 540
68/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
caaagatgaa acagtatttt ttattcaaca aatgaaaatt atagtcattt ctcataacat 600
tccaatgagg ctttttatgc aatggcatgt ttcagtgata aaagttccag ttatgtatct 660
cgacgatgtg acacagtcac agtccctaaa ggaggtcatc acagattttc tcacacaaac 720
tcataaactg tcactctacc tttgccagac tatgaaagta ggcactaaag gaccaggcga 780
taacttacac caagttgcac ctgacctact cctcccagaa caaatcataa agtacttgtg 840
caaatccgaa gaattcatgg acccggcaac atttataaac atgtataata gacccataca 900
catcactctg gattctgagg tggtatggcc tgctccttca cgtttctagg attgagaggg 960
ataatgtgcc catgtctctt aaggagtttg tgctacttaa ataaaaaaaa catttttaaa 1020
g 1021
<210> 68
<211> 4074
<212> DNA
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No:'3434684CB1
<400> 68
ggcggccgag cgaaatataa ctcatatatg gcgaacttgt gcttctagat catgcgtcga 60
gcggcggacg ccagttgtgc gtgcaaacgg gatagacggg tgggccgagg tacaggcccc 120
acggccgccg tctcccgctt ctgcccgcgc agagtccgcg ccatggccgc ctcgccgggc 180
tcgggcagcg ccaacccgcg gaagttcagt gagaagatcg cgctgcacac gcagagacag 240
gccgaggaga cgcgggcctt cgagcagctc atgaccgacc tcaccctgtc gcgggttcaa 300
tttcagaagc ttcagcaact gcgccttaca cagtaccatg gaggatcctt accaaatgtg 360
agccagctgc ggagcaatgc gtcagagttt cagccgtcat ttcaccaagc tgataatgtc 420
ggggaacccg ccatcacggg ctggtggaga ggccatccag gaaccgttcc accccctcca 480
ccgaaggtct ggggacaagc caggtcgaca atttgatggt agtgcttttg gagcccaatt 540
attcctcaca gcctctggat gagagttggc caaggcagca gcctccttgg aaagacgaaa 600
agcatcctgg gttcaggctg acatctgcac ttaacaggac caattctgat tctgctcttc 660
acacgagtgc tctgagtacc aagccccagg acccctatgg aggagggggc cagtcggcct 720
ggcctgcccc atacatgggt ttctgtgatg gtgagaataa tggacatggg aagcatcttt 780
ccactggccc attgacaaga agagaatctg ttaaatgttc cgaagccact gccaaaacaa 840
ctgtgggaga ccacggagat tcagtcctgt caggacgccc tcgatcctgt gatgttggag 900
gtggcaatgc ttttccacat aatggtcaaa aacctaggcc tctcaccctt cttggggacc 960
ttgaacactt ggagggtcat tgccagatct aaccaaaccc cactactcga cacccctgcc 1020
agcctccctg gacaccaccg accaccactt tggcagtatg agtgtgggga atagtgtgaa 1080
caacagccca gcagccccac agtcagactt tcagcttctc ccggcccagg gctcatcttt 1140
gaccaacttc ttcccagatg tgggttttga ccagcagtcc atgaggccag gccctgcctt 1200
tcctcaacag gtgcctctgg tgcaacaagg ttcccgagaa ctgcaggact cttttcattt 1260
gagaccaagc ccgtattcca actgcgggag tctcccgaac accatcctgc cagaagactc 1320
cagcaccagc ctgttcaaag acctcaacag tgcgctggca ggcctgcctg aggtcagcct 1380
gaacgtggac actccatttc cactggaaga ggagctgcag attgaacccc tgagcctgga 1440
cggactcaac atgttaagtg actccagcat gggcctgctg gacccctctg ttgaagagac 1500
gtttcgagct gacagactgt gaacagaagg cagtggaaca gaagaatgtt tttctgcaac 1560
agccaaaata gaatggaata gaatgaagcc agctgatacc acgggctttc gttatcttga 1620
catagaagga agcaatgcca cggctccagg gtttcagatg agatcccatc tcagacactg 1680
tggcttcctc cagatcacac agctttgtac tgcctctccc gcctgtggcc aaagtcgtgt 1740
tgcagcaggc aggctgcttg gagcttccca tgaactggaa agctcacctc cactgcatct 1800
ttttactggc catccagtca gccgatgtgt aagagtagga aatactgtgt cactggaggc 1860
ctccgtagca ttgtgtagtg tgctcagaac cactgatctc cgtccgcacc gaaggcgggc 1920
ccggagtggg aggctcggcc tggggcggcg gcaccggaga gggcacctcg atgcctgctc 1980
tgacctgacc cagagggcga ggccctccag cgggggacat tcccaggctg agtggacccc 2040
acggctctct cccacgcctg gattacgaca tgaagttttt accacaagcc cgagggcagg 2100
cttgagttag gcagactgaa ggctaatttt cattttctcc cagctggttt ctgctgcttc 2160
agaaaagtac accttttcct tatggaccag aggaagagga agaccatttt atcagtcact 2220
gaaaagagtc ccctggcact gatgagcctg aagagaactg tgctcctcct cgggggctgg 2280
ggaaggaaga cagggactgc aggtgtctca tactcagtgg cctccagaca aactccagac 2340
aagcacagac ctccccacta agagcagcca gagggagctg gtgaggccct aaccccaccc 2400
accgagcaac tgagcttccc catccctccc cagagctgtg tctctgtggg ctgggagtct 2460
aatgtcaccc ccctagaccg taagctcctt gagggcaggg acagtgcctt atcattgttg 2520
aacccgtagc acctaacacg tgcgtggcag cacagtcgtg tgctggagtg agtgctgcag 2580
69/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
aaccgtgcgt gcagcgcatg atgaatgagt gcgtccgcca tgccgtaagg caggctcacc 2640
tgtagctatc cccttccctg ccagatcttc tcagagcttt agcttttcta gcactcgtgc 2700
ctatggtgaa gcatgcactt taatatgctt ttaacactag gtgaccaaat cacagtgaag 2760
ccgggcactg cattctcctt gggctgtgct ccacgcgggt gggtgggagc tgtcttctga 2820
gtacatccgg aagggctgag caggtgagtg ccctgagcat cctggctggg CCCCaCCCag 2880
gaagatcttc cttctcagat ccacgtttgg ctctaaattg cttcaagtag agattcattc 2940
tttgaggttg aaaaaatagt ttatagtaaa acgaaggcct aattcatgga agcatcatta 3000
gtcatcaata ccttctcata aaatagatgg gccagatttc caccaccgcc tgcctcctct 3060
aacttgggtg atgccagtag gtttgaaggg ggcagagcac tgcaggggga ggggggggtc 3120
taggctgtga gaggacaggg tggagaggag gagaaccctg aaggagagac ggaagatgcc 3180
aggacctttg cttggacagc cattgccctt ccaggagacc ctggagtgta ctgagggttg 3240
ctggactgtt ccacccagag gagcaaggct gtacaatgag ggtctgaatc tggcacacct 3300
gtcccttatg taaaaggagt cgtggtcaca agaccctggg ctggttagcc tctcccgacc 3360
tcattccatt tctatcttct gaccttgcct ctcatcttta aaataaccct catggggtgc 3420
ccctccacct tcctctggaa tccaagtatt cctgtttcac atttgcccca aatctttgct 3480
gtggaattgg gaaatcaaac cagagtcctc ctcgcctgat ttccagctca ggaagggcct 3540
gctggcctgc cctgttccca gttacacttt cagatccctt tgtgctcaag atctcagagg 3600
gggtggcttt ttgttaaaga gccttcagtc gcaatgctac ccagcacccc atgtgccaaa 3660
agaaaccagc tcctgtgtca aagggcttcc aaacctgatc tcactctcaa caggcgatgg 3720
tgctgatgtt tcaagaattg tgtttttata aaacagaggt ctcagcatag tcactcttca 3780
catagtgcct tacccatggg tatcgtcata tccgggtccc agttcagttt gtctgcacag 3840
cagccaccct gcctggcaac agagacccca agactacaca gtgaacccta ctgccccaaa 3900
ggcgttctcc aggtgacttg tgaaaacaga cctccgggga agtgatttat gggggtgtac 3960
actgggggca tatggtttag cactgaattc aatttgtctt aggtctatga gtgagtcgat 4020
cttttcttgt gaaggtttgg gctcgttaca accactctgc ctagaggtgt gggg 4074
<210> 69
<211> 2551
<212> DNA
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 5134056CB1
<400> 69
gcgctaggca gcttcagccg gaccgggtag gggtcctcgc tcgctagctt gctgtttctc 60
ggagaagctc ccgagtgtcc ggcctagagg ccatgagaag gcagtggggg tctgccatga 120
gggcggccga gcaggcgggc tgcatggtga gcgcctcccg ggccggacag cccgaggcgg 180
gcccgtggag ctgcagcggg gtaatcctga gccgtagccc gggcctggtg ctttgccacg 240
ggggcatctt cgtccccttc ctgcgagctg gcagcgaagt cctgaccgcg gccggcgccg 300
tcttcctgcc tggcgacagt tgcagggacg acctgcgcct gcacgtgcag tgggccccaa 360
cggccgcggg tcccgggggc ggcgcggagc ggggccgccc agggctgtgc acgccccagt 420
gcgcgagcct cgagcccggc ccacctgccc cgtcccgcgg gcgtcccctg cagccccggc 480
ttcctgctga gctgctgctg ctgctgagct gcccggcctt ctgggcccac ttcgcgcgcc 540
tcttcgggga cgaggcagcg gaacagtggc gcttctcgag cgcggcgcgg gatgacgaag 600
tgtcggagga cggggaggcg gatcaactga gagcgctggg ctggtttgcg ctgctgggcg 660
tgcggctagg ccaggaggag gtggaggagg agcgcgggcc agccatggcg gtgtcgcctc 720
tcggggccgt gcccaagggt gcgccattgc tggtctgcgg ctcccctttc ggcgccttct 780
gccccgacat ctttctcaac acgctgagct gcggggtgct cagcaacgtg gccggcccac 840
tgctgcttac cgacgcacgc tgcctgcccg gcaccgaggg cggcggcgtg ttcaccgcgc 900
ggcccgcggg ggcgctggtg gcgctggtgg tggcgccgct ctgttggaag gccggcgaat 960
gggtgggctt cacgctgctc tgcgccgccg cccccctttt ccgcgccgcc cgcgacgcgc 1020
ttcaccgcct gccgcacagc accgctgccc tggccgccct tctgccgcca gaggtgggcg 1080
tcccgtgggg tctgcccctc cgagactccg ggcccctgtg ggcagccgcg gcagtgttgg 1140
tggagtgcgg caccgtatgg ggctccggag tggctgtggc accccgcctt gtagtgacct 1200
gtcggcacgt gtcccctcgg gaagcagcca gggtcctggt~gcgctccacc acccccaaga 1260
gtgtggccat ctggggccgt gtggtatttg ccactcagga gacatgtccc tatgacatag 1320
cagtggtgag cctggaggag gacctggatg atgtccccat ccctgtgccc gctgagcact 1380
tccatgaagg cgaggctgtg agtgtggtgg gctttggcgt ctttggccag tcttgcgggc 1440
cctcggtgac ctcaggcatc ctttcggctg tggtgcaggt gaatggcacg cccgtaatgc 1500
tgcagaccac gtgtgctgtg cacagcggct ccagtggggg acccctcttc tccaaccact 1560
caggaaacct ccttggcata atcaccagca acacccggga caataatacg ggggccacct 1620
70/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
acccccacct gaacttcagc attcccatca cggtgctcca gccggccctg cagcagtaca 1680
gccagaccca agacctaggt ggcctccgtg agctggaccg cgctgctgag ccagtcaggg 1740
tggtgtggcg gttgcagcgg cccctggcag aggccccgcg gagcaagctc tgaggctgtg 1800
ttaccacctt tggaaagaag agtgaccttt ttctgctgta ggaagtgatg ttgaggtgac 1860
ggtggcctca ggattcaggg cccagcccct gcaggggccc aggctgcctc tcatctccac 1920
ccactgactg cagactgggc tttgggctct ggggcaaact tctcttcagc cccatggatc 1980
cttaacctgg cagcccgttt tggggtgctt tcttgagccc ccagttctct gtcccctagc 2040
actagactca gctgtattgt ttttccttct ggggagccca ctccaactgc acagaagttc 2100
tgggcctgac aggtagattc cagctggaag gcaggcccgt gcctggtttt gcgtctgttc 2160
ccctgagggc catcgtcatc ctggagcttc aatggggcct tggctcctgt ctgcctctca 2220
gtcagagtca gggctgacaa aggactcagc ttccttagca tctcagcaga aaccttgctc 2280
tgaagaccag agacagaagg gacagaaaca ggagtgcctc ctgctgtgcc aggcccatgg 2340
gcagtgcagg cagatccctg aaggtcagca ctcctgggtc ttcatatgcc aacaggggcg 2400
ctcttgacac tgtgccttca ttttccagcc cacagcctgg gtctcaggga tcttgagggg 2460
tagaacatgt ctggttgggg cttgggaata aacatgatct attgaaaaac ctctgtcaaa 2520
aaaaacaaaa aaaaacaaca aaacagaaaa t 2551
<210> 70
<211> 1115
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 5281724CB1
<400> 70
ggcgccgcag tgtctacacc cgcaaaaacg cccgctttga cttagagccc tccgcatatc 60
cttcccctgg ctggggcagc caaggcgcgg aggacagctc cgaggccaga tgtttactgc 120
gcagatgccc gactttacac cggggtccgg gctgtgccgg gcgctggggg aagcgcccac 180
cttccagaga gcgaaatcat ggagccttcc aagaccttca tgagaaacct gccaatcaca 240
ccaggctata gcggctttgt gccattcctc agctgccaag gaatgtccaa ggaggatgac 300
atgaaccact gtgtgaaaac cttccaggag aaaacacagc gctataaaga acagctgcgg 360
gaattgtgct gcgcagtggc cactgccccg aaactgaaac ctgtcaactc cgaggagacg 420
gtcctgcagg ccctgcacca gtacaatctg cagtaccacc ccctgatcct ggaatgcaaa 480
tatgtaaaga aacctctcca ggagcccccg atccctggct gggcaggcta cctgccgaga 540
gccaaggtca Ctgaatttgg ctgtggcacg agatacactg tcatggccaa aaactgctac 600
aaggacttcc tggagatcac ggagagggcc aagaaggcac atctgaaacc atatgaagaa 660
atatatggag ttagctccac aaaaacttct gctccgtctc caaaagtttt gcagcatgaa 720
gagctgctgc caaaatatcc cgatttttct attccagatg gaagctgccc tgcccttgga 780
aggcccctga gagaggaccc caaaactccg ctgacatgtg gctgtgctca gaggccaagt 840
ataccatgca gtgggaagat gtatctagag ccactgtcct ccgcaaagta tgcagaaggc 900
tagaagcgca gagtctccca aggaggtgaa ctttaagtgg ggcttccaaa acctgccatt 960
ctcatgttgg aatcacgccc agtgagcaat aaagaaattt agtaacaaga attttttatc 1020
tgccgcctgc atcctgagtg gttcccggtt gcatgtcatt aatgataaag gccgttcttt 1080
gtcatgtcgg aataaagagg gtgcttctcc gcaaa 1115
<210> 71
<211> 1334
<212> DNA
<213> Homo Sapiens
<220> ..
<221> misc_feature
<223> Incyte ID No: 7502391CB1 ,
<400> 71
cgcgagccgg gctgtcgggt gtgttttgct ctccatcctc cgtcgtctct gcagcactcc 60
gggttctcct ccagagcgct agtcccagga gctcggaatg ttcgtggaac ttaataacct 120
gcttaacacc acccccgaca gggcggagca ggggaaactg actctactct gtgatgccaa 180
gacagatggg agtttccttg tacaccactt tctctccttc tatctcaaag ctaattgtaa 240
agtctgcttt gtggcactca tccagtcctt cagccactac agtatcgtgg gacagaagct 300
gggtgtcagc ctgaccatgg cgcgggagcg tgggcagctt gtgttccttg agggactcaa 360
71/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
gtctgcagtg gacgtcgtct tccaggctca aaaggagcca caccccctgc agtttctcag 420
ggaggctaat gctgggaact tgaaaccatt gtttgagttt gtacgggagg ccctgaagcc 480
agtagacagt ggagaggctc ggtggacgta cccggtgctg ttggtggacg acctcagtgt 540
gctcctgagc ctgggcatgg gggcggtggc tgtgctagac ttcattcact actgcagagc 600
caccgtgtgc tgggaactaa agggaaacat ggtggtcctt gtgcacgaca gtggagatgc 660
ggaggatgag gagaatgaca tcctgctgaa tggcctcagt catcagagcc atctgatact 720
gcgggctgag ggcctggcca ctggcttctg cagggatgtg cacgggcagc tgaggatcct 780
gtggaggaga ccatcgcagc ccgcagtcca ccgggatcag agcttcactt accagtataa 840
gatacaggac aaaaggcgtg tccttttttg ccaaaggaat gtctcctgct gttctgtgac 900
ctgatttcgg agcagctgaa gctacatagg actgtttttg gacgtggaag atagagcaac 960
atagcaagaa tgggtctttc tcctctgtag taatatttca ggctggaccg gcgactccac 1020
tgtgaccaga gggttgagtg ctgcagtgat ggcatgcctt ggctgcctgg gccctgttca 1080
gaaaacacaa gggaccacaa tcctgccttt gctgagagag aggctggatg ctagacccaa 1140
gtgaaagggg tcctttggag cctttgttta aatatgcctt agccccagct gcccattttt 1200
ggttgacaag cctttcagag ccagagtggg tatagatgtg ccagccagga gatggcaccg 1260
gatggcaggt gtgcaaggtg acaactagga taatcatggc tggaataaag taagtttcca 1320
cactggaaaa aaaa 1334
<210> 72
<211> 2387
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7502544CB1
<400> 72
cgctcgcgcc ggaccggaaa gccggggaag tggccgagga gggagggctg cgagccatgg 60
cgaccaagac ggcgggcgtg gggcggtggg aggtagtgaa gaagggtcgg cggcctgggg 120
tcggcgccgg cgccggcggc cgaggaggcg gcaggaaccg cagggcgctc ggggaagcaa 180
acggagtgtg gaaatacgac ctgacccctg caatccagac cacaagcacc ctttatgagc 240
ggggctttga gaatatcatg aagcggcaga ataaggagca ggtcccaccc Cctgctgtgg 300
aacctaagaa accagggaac aagaagcagc caaagaaggt ggcaactcct cccaaccaaa 360
accagaagca gggccgcttc cgcagcctgg aggaagcact gaaagctctg gatgtggcag 420
acctgcagaa ggaactggac aagagccaga gtgtgttctc tggaaaccca tccatatggt 480
tgaaggacct ggccagctat ctcaactaca agctacaagc tcctctaagt gaacccacgc 540
tgagccagca tactcatggc ctatgggcct tagattatcc ctacagcctg gtgagccggg 600
agctacgtgg gatcatccga gggctgctgg cgaaggcagc agggtctctg gagctctttt 660
ttgaccactg tctgttcacc atgttgcaag agctggataa gacaccaggg gagtcactac 720
atggttaccg catctgtatc caggccatcc tgcaagacaa gcccaagatt gccacggcaa 780
acctaggcaa gttcctggaa ctgctgaggt cccaccagag ccgaccagca aagtgtctca 840
ccatcatgtg ggccctgggt caagcaggtt ttgccaacct caccgaggga ctgaaagtgt 900
ggctggggat catgctgcct gtgctgggca tcaagtctct gtctcccttt gccatcacat 960
acctggatcg gctgctcctg atgcatccca accttaccaa gggcttcggc atgattggcc 1020
Ccaaggactt cttcccactt ctggactttg cctatatgcc gaacaactcc ctgacaccca 1080
gcctgcagga gcagctgtgt cagctctacc cccgactgaa aatgctggca tttggagcaa 1140
agccggattc caccctgcat acctacttcc cttctttcct gtccagagcc acccctagct 1200
gtccccctga gatgaagaaa gagctcctga gcagcctgac tgagtgcctg acggtggacc 1260
ccctcagtgc cagcgtctgg aggcagctgt accctaagca cctgtcacag tccagccttc 1320
tgctggagca cttgctcagc tcctgggagc agattcccaa gaaggtacag aagtctttgc 1380
aagaaaccat tcagtccctc aagcttacca accaggagct gctgaggaag ggtagcagta 1440
acaaccagga tgtcgtcacc tgtgacatgg cctgcaaggg cctgttgcag caggttcagg 1500
gtcctcggct gccctggacg cggctcctcc tgttgctgct ggtcttcgct gtaggcttcc 1560
tgtgccatga cctccggtca cacagctcct tccaggcctc ccttactggc cggttgcttc 1620
gatcatctgg cttcttacct gctagccaac aagcgtgtgc caagctctac tcctacagtc 1680
tgcaaggcta cagctggctg ggggagacac tgccgctctg gggctcccac ctgctcaccg 1740
tggtgcggcc cagcttgcag ctggcctggg ctcacaccaa tgccacagtc agcttccttt 1800
ctgcccactg tgcctctcac cttgcgtggt ttggtgacag tctcaccagt ctctctcaga 1860
ggctacagat ccagctcccc gattccgtga atcagctact ccgctatctg agagagctgc 1920
ccctgctttt ccaccagaat gtgctgctgc cactgtggca cctcttgctt gaggccctgg 1980
cctgggccca ggagcactgc catgaggcat gcagaggtga ggtgacctgg gactgcatga 2040
agacacagct cagtgaggct gtccactgga cctggctttg cctacaggac attacagtgg 2100
72/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
ctttcttgga ctgggcactt gccctgatat cccagcagta ggccctgcct tcctggccac 2160
tgatttctgc atgggtagac catccaagac tgcagcgggt agaaggtggc agttcttcat 2220
gggagtcttt ttaacttggt gcctgagttc tctcctaggc aagtggccag ttgcctccgc 2280
ctcagttctt ccatctttgg tggggacagg gcccagcagc atctcagcct cctacccaca 2340
attccactga acacttttct ggccctactg cacatggccc ccagcct 2387
<210> 73
<211> 2919
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 2858465CB1
<400> 73
gtagatgcga tggcgccgat tccaaagact gtggggcgga tcaagctaga ctgctctcta 60
cggcccagct gcccactgga ggtcgctgct gcacccaaac tttgcaagga attcggtcca 120
gaggattacg gcgaagagga catagtggat tttcttcgac ggcttgtgga gagtgatccc 180
cagggcctgc accggatcca tgtggatggg agcagcgggc ggctgcagct gtggcaccat 240
gattacctcc tgggccactt ggatgatgaa gggaaatcaa ctggacagag tgacaggggc 300
aagggggctg agggactggg cacctactgt ggtctccgca agtccttcct gtatcctccc 360
caagagtctg agccctgccc tcaaagcccc tctgcctctg ccaccttccc cagtgtctca 420
gacagcctgc ttcaggtggc catgccccag aagctcctgg tgacagaaga ggaagccaat 480
cgcctggctg aggagctggt ggctgaggag gagcgcatga aacagaaagc agagaaaaag 540
cgactcaaga agaagcgtca aaaggaacgg aagcgacagg agcgtttgga gcagtactgt 600
ggggagccca aggccagcac tacctcagat ggagatgaga gccccccatc cagccctgga 660
aacccagttc agggacagtg tggtgaagaa gaggactcac tggatctatc tagcactttt 720
gtgtctctgg ctttgcgcaa ggttggggat tggcccctca gtgcccgcag agagaaggga 780
ctgaaccagg agccccaagg caggggtctg gccctccaga agatgggtca agaggaagag 840
agccctccaa gagaggagag gccccagcag agtccaaagg tacaggcatc tccgggactg 900
ctggcagctg ccttacaaca gagccaggaa ctggcaaagt tgggtaccag ctttgctcaa 960
aatggtttct accatgaggc cgtggtcctc ttcacccagg ccttgaagct caacccccag 1020
gaccaccggt tatttggaaa tcgttccttc tgccatgagc ggttgggtca gccagcgtgg 1080
gccctggctg atgcccaggt ggcccttacc ctacggcctg gctggccccg gggcctcttc 1140
cgcctgggca aggccttgat gggactacag cgcttcagag aggcagctgc tgtgtttcag 1200
gaaactctga gaggtgggtc ccagcctgac gcagcccgag agctccgctc ttgccttctc 1260
cacctcacac tgcagggtca gcgaggagga atctgtgcac cacctctgtc acctggggcc 1320
Ctccagccac ttccccatgc tgagctggca ccctcaggcc taccttccct caggtgccct 1380
cgaagcactg ctttgaggtc ccctggcctg tctccactct tgcattatcc ttcatgtcac 1440
cgaagccacc ccaaccagcc cctctcccgg actcagagta gaaggcccca tcctctcaag 1500
ccccaggacc cttcaaaggg ctgggacatc ctgggacttg ggctccagca tctgtctcag 1560
gccagatgag ggggcaccgg tccctcatag ggcagggcca tgtatatatc ccttggtggg 1620
ggacatagtg tggtgacagt tcactgcata ttttgagacc ttattctcta gatccatagt 1680
taatgatgcc ctggcagtca ttcctcttgc catggggaag cttctgatga gagaaaggag 1740
ccccacatcc actgaaacat cctttggttc tcaagcttct tctggaggca gtaaggaaaa 1800
ataaaaccca ccaaggctca agaagggaac tatagaaaag ttcaggtttt taggctatag 1860
cagagacagt gagaaagcat ctgggccttt Ctcttcctct tggtccaggg gacctcattc 1920
accaactaga gcttggtgta caggaacggg gtcacagtgc tgagggggct tgagtcccac 1980
ctttcagctt gatggatgct cacctcttct cagccccagc tcgtgccctg tttttctagc 2040
catagccccc agattactca cagctcctca tgccatttcc tgtccagatt gctatgtatg 2100
actctgacct ctcttgtcca gtggtctggt gctcacctcc tctcactgct agaatattca 2160
ccaagggttt gcatttggga agtcccttac cagctcctgc ttagagctgg tagggccata 2220
catgtccaca ctcccaactg gtggctctcc cgctgaatgg ggcctcagca ggtgcccaag 2280
ctgctacaac cttggccact ctgtttctcc accccagcac tgggcatggt aattagcctt 2340
tccccatgtt aatttattca gttttttcaa gggtcaactg aattccccac ttcctgggta 2400
agaagcatga tctcctttta atttcacgtc taagatcctg gcagcttccc ctagctggtt 2460
cctctgtagt cctgctggga ctgtcagctc atttaaatgt gggtctgcag aaggctttag 2520
gtctccccca acccccttac ctttcacaga ggaacctttc atcaggataa atgattattg 2580
ctgccctgtg ggtcttgctc aatactgttc atacctggag agagaaggta ttgaaacatc 2640
tcctttatgt gtgactttcc caaattttta aaaattgttt atggtttagg ccccttaaat 2700
actgtgtagc aggatgaagt ctaccattac cagctgggtc accttggatg ggtctgtcaa 2760
catctaagcc tcagttccct cacctgtaaa aatgagggta gtccctacct cataagggat 2820
73/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
attgtgagga tggaaagcga aagtgtgaga aaatacctcc caagtgcctg gtacatagtg 2880
ggtgctaaat aaaccacttt ttgtctgcaa aaaaaaaaa 2919
<210> 74
<211> 1414
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7503455CB1
<400> 74
ggcgaagagg ggcgcaagct cattgcgttt tgagtctcgg gacccctgtt ggagagacta 60
tggcgctcaa caagaatcac tcggagggcg gcggagtgat cgtcaataac accgagagca 120
tcctaatgtc ctatgatcac gtggaactca cattcaatga catgaagaac gtgccagaag 180
ccttcaaagg gaccaagaaa ggcactgtct accttacccc ttaccgggtc atctttctgt 240
ccaagggcaa ggatgccatg cagtccttca tgatgccatt ttatctcatg aaagactgtg 300
agatcaagca gcccgtattt ggtgcaaact acatcaaggg aacagtgaag gcggaagcgg 360
gaggtggctg ggaaggctct gcttcctaca agttgacttt cacggcaggg ggcgccattg 420
agttcggaca gcggatgctc caggtggcat ctcaagagtt ctatccagga ccccccatga 480
tggacggggc Catgggatac gtgcagcccc caccaccgcc ctaccctggg cccatggaac 540
ctccggtcag cggccccgat gtcccctcca ctcctgcagc cgaagccaag gccgcagaag 600
cagccgccag cgcctattac aacccaggca atcctcacaa cgtctacatg cccacgagcc 660
agccgccgcc acctccctac tacccaccgg aagataagaa gacccagtag gccctcctgc 720
Ctccctgcct cccaccctca tctctctacc ctacccctcc catcggggct gtgctggggc 780
ttggggaggg gagggggcgc Cttgttctcc ctccaggtct gatcataaac aattaccagg 840
aactagcatt gtgggacatt agggcccccg gcctcgggag aggtgccgcc cagcttccca 900
tgccagcccg gagcccacag tgctgcccag cgtacctccc tcaccgtctg gggctcttct 960
gggagcacgg agcatcccct gttcctgttt cactctcagc ttctcccctc gaagggactc 1020
tctggccacc tcctccaccg cagtccagct ccctcagtct ggcacccact gctacactca 1080
gcctcatgag ccacttcaga ccagccaggt gtcttcccgg gccctgccag accctgctca 1140
cattccctct gctggtctgt gctggtctca gaaggccacc gcgcccgcat tccactcagc 1200
cagggtccag ctgcagcccc cgccaccctt ccttcccttc cctgtcctgg gtcatgttgt 1260
tgccaccctg tgtgactttt gaagctgtaa aatgagcttc cagggcttgg ggtggcgtcg 1320
gggcagggcc gccgaggctg gggaggaagc ccttctgcct tttgctggtg tttctggaat 1380
ttgctttccc tcacctctca cttccttcta gaag 1414
<210> 75
<211> 672
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7503479CB1
<400> 75
ggcgggtcac gtgacgcggt gcctggcgcc gagcctccca agatggcggt gtgcatcgcg 60
gtgattgcca aggagaatta ccccctctac attcgcagca cccctacgga gaacgagctg 120
aagttccact acatggtgca cacatctctg gacgtggtgg atgagaagat ctccgcaatg 180
gggaaggccc tggtcgacca gagggagctg tacctgggcc tgctctaccc cacggaggac 240
tacaagatgt tccggaagct acacaactcc tacacagacg tgatgtgcaa ccccttctac 300
aacccggggg accgcatcca gtccagcagg gcctttgata acatggtgac gtcgatgatg 360
atacaggtgt gctgagtgag ctgtgctgcc agccatcgca gaggagcccg cgcacgactg 420
tggtggggcc gtcggtctgt tctggttgcc tcttcctgaa tgggacgcct ggggctttca 480
gggcaggcag ctgtgcatgt tctctcaact aaaggtcttg tgagaggaga tttggctttt 540
tccttccgtg tcagccaagg acttaattaa gaagaattca actaaggact tttctggggt 600
tgtgggcaga ggttgggatc agatggcgca cgtagcctgt ctcagttgcc caaaggggca 660
gagcagggtg ca ~ 672
<210> 76
<211> 3056
74/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<212> DNA
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7218127CB1
<400> 76
ccggtcgggg gcggggccgg gctcggcttc tcttagggac ccggcactcg tcgccctcag 60
tccggctcag ctggggctgg gcgccgtggg tctggcggtt ccgtagcggt cccagcgtct 120
gtcccgccgg ccgggcggtc gcggcacagc cgcgggaagg tgtcggaggg cggttccgcc 180
gcgcggcggg cgcgccgccc acatggcggc catcagggcg ctgcaacagt ggtgccggca 240
gcagtgcgag ggctaccgcg acgtgaatat ctgcaacatg accacgtcgt tccgcgacgg 300
cctggctttc tgcgccatcc tgcaccgcca ccggcccgac ctcataaact tcagtgctct 360
caagaaggaa aatatttatg aaaacaataa actggccttc cgcgtggccg aggagcactt 420
gggcatccca gccttgctgg atgccgagga catggtggcc ttgaaggtgc ctgaccggct 480
gagcatcttg acctacgtgt cccagtatta caactacttc cacggccgct cccccattgg 540
gggcatggca ggcgtgaaga gggcctcgga ggactctgag gaggagccgt cagggaagaa 600
ggctccagtc caggcggcca agctgccctc gcccgcccca gcccggaagc ctccactatc 660
tccagcccag acaaaccctg tggtccagag gaggaatgag ggtgcagggg gcccgccccc 720
caagactgac caggcattgg cgggcagctt ggtcagcagc acctgcgggt ctgcggcaag 780
cacgtgcacc tggtacagcg gcacctggcc gacgggaggc tttaccaccg gagctgcttc 840
aggtgtaagc agtgctcctg cacgctgcac tcgggggcct acaaggccac aggagagccg 900
ggcaccttcg tctgcaccag ccacctcccc gcagccgcct ctgcaagccc caagttgacg 960
ggtctggtcc cccgacagcc aggggccatg ggtgtggatt ccaggacctc ctgttcccca 1020
cagaaggccc aggaggcaaa caaggccaga ccgtcggcct gggagcctgc tgcgggcaac 1080
tcgcctgcca gggcttccgt tccagctgca Cccaaccctg cagccaccag Cgccacgtcc 1140
gtccacgtga ggagcccagc caggccctct gagagccgcc tggcccccac tcccacggag 1200
gggaaagtcc gccctcgtgt gaccaatagc tccccgatgg gctggtcgtc agctgccccg 1260
tgcacagcag cggctgcctc ccatcccgcc gtgcccccga gtgccccaga ccctcgcccg 1320
gccacacccc agggcggggg agccccccga gtggcagctc ctcaaaccac actcagttca 1380
agctccacat ctgcagccac ggtggacccc ccagcctgga ccccgtccgc ctccaggacc 1440
cagcaggccc ggaataagtt tttccaaaca tcagcagtgc cccccggcac cagcctttct 1500
ggcagaggtc ccaccccgtc acttgttcta tccaaggaca gcagcaagga gcaggcgcgg 1560
aacttcctca agcaggccct ctcagcgctg gaagaggctg gcgctccggc gcctggcagg 1620
ccctccccag ccactgccgc tgttcccagt tctcagccca aaactgaagc accacaagca 1680
agtcccttag ccaagccgtt acagtcctcg tctccccggg tgcttggcct gccttcgagg 1740
atggaaccgc cagccccgct gagcacgagc agtacctctc aggcatccgc gttgcccccg 1800
gcaggcagga ggaacttggc tgaatcctca ggggtcggca gggtgggtgc tggctccagg 1860
ccgaagccag aggccccgat ggcaaagggt aaaagcacca ccttaacgca ggacatgagc 1920
accagcctcc aggaaggcca ggaggacggg ccggcaggat ggagagcgaa tctgaagccc 1980
gtggacagga gaagcccagc tgagaggact ctgaagccca aggaaccacg ggccctggca 2040
gagccgaggg cgggggaggc ccccaggaag gtctcaggca gctttgctgg gagtgtccac 2100
atcaccctga cccccgtgag gcctgacagg accccacgcc cagccagccc aggacccagc 2160
CtCCCagCCa ggtccccctc CCCaCCCCgC CgCaggagaC tggccgtccc tgccagcctc 2220
gacgtttgtg acaactggct tcggccggag ccccctggcc aggaagcccg agtgcagagc 2280
tggaaggagg aggagaagaa acctcacctt cagggcaaac cagggagacc cttgtccccg 2340
gccaatgtcc ctgctctgcc tggcgagacg gtgacctccc cagtcaggct gcaccccgac 2400
tacctctccc cggaggagat acagaggcag ctgcaggaca tcgagaggcg gctggacgcc 2460
ctggagctcc gcggcgtgga gctggagaag cgactgcggg cggccgaggg agatgacgct 2520
gaggatagcc tcatggtgga ctggttctgg ctcattcacg agaagcagct tctgctgaga 2580
caggagtcag agctgatgta caagtccaag gcccagcgtc tggaggagca gcagctggac 2640
atcgagggcg agctgcgccg gctcatggcc aagcccgagg ctctgaagtc actgcaggag 2700
cggcggcggg agcaggagct gctggagcag tacgtgagca ccgtgaacga ccgcagtgac 2760
atcgtggact cgctggacga ggaccggctc cgggaacaag aggaggatca gatgctgcgg 2820
gacatgattg agaagctggg cctccagagg aagaagtcca agttccgctt gtccaagatc 2880
tggtcaccaa aaagcaaaag cagcccctcc cagtagtagc cagtagggcc gtgggctcgg 2940
cccggacctg gcatccggac ttggactccg ggtcaagccc atggccccga gtccacgtcc 3000
gactgccagg tccggagcgc gacgcgtgga aagccgaatt ccagcacact cgggcc 3056
<210> 77
<211> 1891
<212> DNA
75/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 1688943CB1
<400> 77
gtggaagctg gcctggcccc cggagctccc tggagtcggt actgggggct tcgttttgta 60
cgcaccgttt tctctctgtg ctatgggaga tgtcaaggaa tcaaagatgc aaataacacc 120
agaaactcca ggaaggatcc ctgttttaaa tccttttgaa agtcctagtg attattctaa 180
tctccatgaa caaactctcg ccagtccttc tgtttttaaa tcaacaaaat taccaaatag 240
ataaagatgt ggaagacaaa agacaaaaag ccattgaaga gtttttcact aaagatgtca 300
tcgtaccctc tccttggact gatcatgaag ggaaacagct ttcacaatgt cattccagta 360
aatgcactaa cataaatagt gactctccag ttggaaaaaa gctgaccatt cattctgaga 420
aaagcgatgc tgcttgtcag acattgctgt ctcttcctgt ggattttaat ttagaaaata 480
tattaggtga ctattttaga gctgatgaat ttgcagatca atctcctgga aacctcagtt 540
cttcatccct cagaagaaag ctgtttttag atgggaacgg aagcatctcc gactccttac 600
cttcggcttc tcccggaagt cctcacagtg gtgttcaaac atcactagag atgttttatt 660
caatagattt gtctcctgta aagtgtagga gccccttgca gacaccaagt tcggggcagt 720
tttcttctag ccctattcag gctagtgcaa aaaaatacag cttgggaagc ataactagtc 780
cttcgcctat ttcttcaccc actttctcac caattgaatt tcagatagga gagactccac 840
tctcagaaca aaggaagttt actgttcatt ctcctgatgc ttcatctgga acaaattcta 900
atgggataac taatccgtgt atcagaagtc cttatataga tggctgctcg ccaattaaaa 960
attggtctcc tatgagactt cagatgtata gtggtggtac tcagtatcgg acctcagtga 1020
ttcagatacc ttttactctt gagactcaag gtgaagatga ggaagataaa gagaatattc 1080
cttccacaga tgtctcatca cccgccatgg atgctgctgg aatacaccta cggcagttta 1140
gtaatgaggc ttctacccat ggtacacatt tggttgtgac tgccatgtct gttacacaaa 1200
atcagtccag tgcttctgag aaagaattag cactgttgca ggatgttgaa agggagaaag 1260
acaataacac tgtggatatg gttgatccta tagagatagc agatgagacc acttggatta 1320.
aggagccggt tgataatggc agtttaccca tgactgattt tgtaagtggc attgccttca 1380
gtattgaaaa ctctcatatg tgcatgtcac ctcttgctga aagcagtgtc attccttgtg 1440
aaagcagtaa cattcagatg gatagtggct ataatacgca gaattgtgga agcaatatta 1500
tggatacagt tggggcagaa agttactgca aagaaagtga tgcacaaaca tgtgaagttg 1560
agagtaaatc tcaagcattt aatatgaagc aagaccacac aacacagagg tgttggatga 1620
aaacagcaag cccttttcaa tgcagcagtc Catagaatac ctctgtcaga atcaaagact 1680
aagcttaaga gttcctcgca tatatcgttg tgcacaggat caacatgatg gtgactggga 1740
aaaaattact tcaagtaaca tgcttagctt tccctcctta atgtgaaaaa tcaagggctt 1800
actgacatag gaacaacaga aatgctcctg gaacttcaag ttgctgaatt ataagtttat 1860
tttttatcaa taaatatttt tatacttaaa a 1891
<210> 78
<211> 2221
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 2369350CB1
<400> 78
ggtggggccg ccgaggtgta cgtctgtgga gccgagccgc ctccttgcct tctccgcggg 60
ccgctctttt agcctgcgcg gccgtcctcc gagcaggccg ccccagcttt ggagttcctc 120
ctcactgcac ccctgtttct gctcactgag accagcttgt gggaaggagc gccccccgcc 180
tcactcacca accccttggc tggcagcaca gcagactctg gggcggcctc gggttcctgg 240
ggtgatagtc cagcttctgg tgtaccagag gaagaagaaa ggcgtgtccc ccaaaggctg 300
tccttgcagt agctggaagc cagatgagta acgaaagagg ctttgaaaat gtagaactgg 360
gagtcatagg aaaaaagaag aaagtcccaa ggagagtcat ccactttgtt agtggtgaaa 420
caatggaaga atatagcaca gatgaagacg aagttgatgg cctggagaag aaagatgttt 480
tgcctactgt tgatccgaca aaacttacct ggggtcccta cttatggttt tacatgcttc 540
gggctgctac atcaactctc tcagtgtatg acttccttgg agagaagatt gcatctgttt 600
tgggtatcag caccccaaag taccaatatg ccattgatga atattatcgg atgaagaagg 660
aggaagaaga agaagaagaa gaaaacagga tgtctgaaga agcagaaaaa caatatcaac 720
agaataaatt gcagactgat tccattgttc agacagatca accagagaca gtgatatcca 780
76/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
gctcatttgt gaatgtcaat tttgaaatgg agggagacag tgaagtaatt atggaaagca 840
agcaaatcca gtctctgtcc caccataaaa tgaaatgact atcaagcttc caactcttaa 900
gttttttttt tttaatacaa aactttcaca ttctttattc agtgggactt aatacaatta 960
tttatatttt aaattattaa agtatctgga aagggaaaat gttttcttca tttttaggat 1020
ctatctagca aagccagatc tgaaattcag atatttgtac tgtttttact gtgtatagaa 1080
attagtgctt tggttttaaa atgatctttt aaaaaagtta aggacatcct agagccttaa 1140
tagttaagaa gagttaaatt atcaagccta tttgtgcatt tgcttttttt gaaaaaggta 1200
agttgctgat taagtctaat tggaattgat aattccatag tcttagatta aaatgaggat 1260
attttctcct agattttctc atgttatgcc atgcatttat atatctaacc attaatttca 1320
cactaaggat gcttcaccat ataataaaag gagcaagatg gaagcacttt gaattttctt 1380
tcattgagaa taactgtttt atgtaagaat ctgtatttat aacaccagat attaagatag 1440
gcttccattt tttaatgcaa gccacttact taatcttgta ttctttttca ggactcaaat 1500
aactagcttt gaacataata ttaaaacact acttatagaa tagatttatt aatgttaata 1560
cctagtgaat atccatgtgg catcctggtt atgttatcgg ttcagcgtta atcctataga 1620
aaagtggttt ggaggggatt gggggatagt gggacaggta tagatttaat ccatcaggag 1680
caattagata ttgtataagg tgcaatgata gcctaatgaa attacccgtc attcatoatt 1740
tagaagtagc aacagtgaag actggacagt ttacttgaat ctggttggcc actcctctac 1800
ctacttggtt atttgtaaac cttacaaatg tatatattgt gaagctaatt ttgaaaatat 1860
tcctaaatat ggccaggtac ggtgctcaca cctgtaatcc caaagtgctg ggattacagg 1920
catgagccac cgcacctggc cttccctgct tcctctctag aatccaatta gggatgtttg 1980
ttactactca tattgattaa aacagttaac aaactttttt Ctttttaaaa tgtgagatca 2040
gtgaactctg gttttaagat aatctgaaac aaggtccttg ggagtaatta aaattggtca 2100
cattctgtaa agcacattct gtttaggaat caacttatct caaattgtaa ctcggggcct 2160
aactatatga gatggctgaa aaaataccac atcgtctgtt ttcactaggt gatgccaaaa 2220
t 2221
<210> 79
<211> 2039
<212> DNA
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 2722979CB1
<400> 79
tgacccggtc caaggcggaa aaagtgcggc cgcccactgt gccagtgccg caggtggata 60
ttgtgcctgg gcggctcagt gaggccgagt ggatggcgct tacagccctc gaggagggcg 120
aggacgtcgt aggggacatc ttggccgact tgctggctcg agtcatggac tctgctttca 180
aagtctacct gactcagcag gtgggccggg atccgggtcc ttcagactcg tctccctctc 240
ccgcccctcc ctgccgacct gagatcctct ctcgcctccg cagtgcattc cattcaccat 300
cagccaggcc cgggaggcca tgctgcagat caccgagtgg cgcttcctgg cccgggacga 360
gggagaatct gcagtagctg aggaccccac atggggtgag gacgaggagc cttcggcatg 420
cacgacggac tcctgggctc agggttcagt gcccgtgctg cacgcgtcca cctcggaggg 480
cctggagaac ttccaaggcg aagtacactc ctcaggagcc tctccggact cctctgccat 540
tgctcctgct ctcccctttc cgacatctca ctgcccgagt gcatttcccc aggaccctgg 600
gggcgtggac cggatccctt taggaaggtc gtggatgggt cgaggctccc aggagcagat 660
ggaatcttgg gagccttctc cgcagctgag agtcacgtcg gcccctcctc ccacatcaga 720
gctgtttcag gaggcagggc ccggaggtcc tgtagaggaa gcggacggcc agtctagagg 780
cctctcctcg gccgggtcct tgagcgcgag cttccaactg tcggtggagg aggcgcctgc 840
cgacgatgcc gacccttctc tggatccgta cctggtagcc agcccccagg cctcaactgg 900
gaggggacac cccctcggct tccatttgtc gttggaagac ctctactgtt gcatgcctca 960
actggacgcg gctggggatc ggctggaact caggtcagag ggggtgccct gcatcgcctc 1020
gggcgtgttg gtgtcctacc cctctgtggg CggCgCCaCC CgCCCCtCCg cgtcctgcca 1080
gcagcagcgg gccgggcact cggatgtgcg gctgagcgcc caccaccaca ggatgcgccg 1140
caaggcggcc gtgaaacgcc tggaccctgc gaggctcccg tgccactggg tgcgccctct 1200
ggctgaggtc ctggtcccag actctcaaac acgccccttg gaagcctacc gcggacgcca 1260
gcggggcgag aagaccaagg cccgggccga accccaagcc ctcggccccg gcacccgtgt 1320
ctccccggca gcgttcttcc ctctccggcc aggcattcct ttccgtgact tggactcggg 1380
ccccgcactc ctgttcccca ctttaaattt aggcctatcg tcgccatccc tcgagtcaaa 1440
gctgccactc ccaaactcca ggatccgctt cctcaccaca cacccggtgc tccctgatgt 1500
ggcccgcagc cgcagcccca agctgtggcc cagtgtcagg tggcccagcg gttgggaggg 1560
gaaggccgag ctgctgggcg agctgtgggc tggccggacc cgcgtgcctc cacagggtct 1620
77/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
ggagctggca gacagggagg gccaggatcc tggcagatgg cctcgaacca cacccccggt 1680
ccttgaagcc acttcccagg tgatgtggaa gcccgtgttg ctgccagaag ccctgaagct 1740
ggcccctggt gtgagcatgt ggaaccggag cacccaggtg ttgctcagct ctggtgtgcc 1800
tgaacaagag gacaaagaag gtagcacctt tcctcccgtt gagcaacatc ccatccagac 1860
aggtgcccca aagcccaggt gaccgtagca cagctaatga agaactcagc ccccaaagtg 1920
tggtcacgct cctctaagcc tgctgcctcc ctctgatccc tgagcctctg gcagtagtaa 1980
ctggtccctc cctctgctag ccagaaataa acacctgagt tgccttagga actaaaaaa 2039
<210> 80
<211> 1254
<212> DNA
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 60140470CB1
<400> 80
agcggctgag cctgcgcttc ggaagcatgg atgtgcgcct gcgctgcgct agggcgcggc 60
gggcggtttg aattttgctt acagagtccc gtctcaccat cctgggcttc caacggagac 120
tgcggtatcc gcggctggag acccagcggc gagtagcctt ttgctcccgg acggacttga 180
gaggcttaaa ggatggcctc gtcagatctg gaacaattat gctctcatgt taatgaaaag 240
attggcaata ttaagaaaac cttatcatta agaaactgtg gccaggaacc taccttgaaa 300
actgtattaa ataaaatagg agatgagatc attgtaataa atgaacttct aaataaattg 360
gaattggaaa ttcagtatca agaacaaacc aacaattcac tcaaggaact ctgtgaatct 420
cttgaagaag attacaaaga catagaacat cttaaagaaa acgttccttc ccatttgcct 480
caagtaacag taacccagag ctgtgttaag ggatcagatc ttgatcctga agaaccaatc 540
aaagttgaag aacctgaacc Cgtaaagaag cctcccaaag agcaaagaag tattaaggaa 600
atgccattta taacttgtga tgagttcaat ggtgttcctt cgtacatgaa atcccgctta 660
acctataatc aaattaatga tgttattaaa gaaatcaaca aggcagtaat tagtaaatat 720
aaaatcctac atcagccaaa aaagtctatg aattctgtga ccagaaatct ctatcacaga 780
tttattgatg aagaaacgaa ggataccaaa ggtcgttatt ttatagtgga agctgacata 840
aaggagttca caactttgaa agctgacaag aagtttcatg tgttactgaa tattttacga 900
cactgccgga ggctatcaga ggtccgaggg ggaggactta ctcgttatgt tataacctga 960
gtcccttgtg aacttttgaa cataccaaca gggtatagag tatagaggct atttctataa 1020
ttttcttata tataattttt ttaactttta atcttttttg tttccttttt tttttttttt 1080
gagacaggat cttgctttgt cacccagggg cttgctttgt cacgcaggct agagtgcagt 1140
ggcgcaaaca tggctcactg cagcctcaac ctaccaggct caagtgatcc tcccacctca 1200
gccccctgaa tggctgggac tacaagcgtg cgccaccatg cctggctaat ttta 1254
<210> 81
<211> 1879
<212> DNA
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 70623603CB1
<400> 81
tccacctgcc agcggagttt aaagtttccg aggctcagag gaacacaatg acttggatca 60
aacagcctaa atgggaagaa ggacattttt gctgcatcaa ggaagccgtt aaactcctgc 120
taagctaact agctcttttt tatgggtcca tgcacacgac cgaactcctc tttcactgac 180
cagagattat ttctgacaac ccaggatatc ccgaaagctt ggaggcatat ggctggaaaa 240
tgaaacgacc caggacatcg tttctggctg catcattatt ttgtgtcgcg tagtaccaga 300
tgggcagtca gtgagcggcg cagggatgtg aacggacggt tttataatgt gaaaattttc 360
ccttggtaaa gctaaaacag atttaatttc cctctctttt ctttcactac ttccccctct 420
ttattccccc tctgtctgca atatcagtga actcaacttt gcagtgaggt ggccaaaaag 480
agagagaatg aggagatctt gatcatctta gtgtcggagg agtcgcagcg gactgggaac 540
tgcagctgcg accccccgcg tcctgtgcgg atttcagggc tgataccgca taggcggtta 600
tggaaaggac ggtacaccgg agcggcggag gatagagacc ctggcccccg gagaggtctg 660
ctgatttcgc agcagccttc gaagccgtgg ctgcctttca tctgctgcgt tttattacta 720
ttatcgccgt tccggaaaag tcatggaaga cagcccgctg ccagacctca gagacatcga 780
78/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
gctgaagctg gggcgcaaag tacccgagag tctagtgcgc tctctccgtg gggaggagcc 840
ggttcccagg gaaagggaca gggacccctg cggggggagc ggtggtggtg gcggcggcgg 900
cggcggcggc ggcggctgca gtagcggcag cagctactgc agcttccctc cctccttgtc 960
gtcctcctct tcgtcctccc caacctctgg ctccccacga ggtagccact ctagcgccct 1020
ggagaggcta gaaaccaagc ttcacctcct caggcaagag atggttaacc tcagagccac 1080
agacgtcagg ctcatgcgcc agttgcttgt aatcaatgag agcatcgagt ccatcaagtg 1140
gatgatcgaa gaaaaagcca ccattaccag cagaggcagc agcctcagtg gcagcctgtg 1200
cagtttgttg gagagtcaga gcacctcctt acgtggcagc tacaacagcc tacacgatgg 1260
cagtgatggg ctggatggca tttccgtggg aagttatctg gacacgttgg cggatgatgt 1320
cccaggccat cagacccctt cagacttgga ccaattcagt gacagctccc tcatagagga 1380
ctcacaggca ctacacaagc gtcctaaatt ggattctgaa tactactgct ttggctagtg 1440
acagtttttt gcatgggact ggtgtgcaat gaacttgtat ttatccttct tctccgctgc 1500
tatatttttg gtgtgatttt tattttaata agatgacctt tttaaaagaa gctgattttg 1560
aaactgctta atggtattgc tgttgctcct aatacttctc atctgagctg atttattttt 1620
ctctgttaca tctctatttt ttatttatta caatgatttt ctcccttctt ttacagtagc 1680
acaaacaaag tagggggaaa agaataagca ataattatgt ttttgctttt gttttcagag 1740
caatgggtca gggattacaa gaaaaacttt gctaaatttt acaataaacc aaagtctgat 1800
aacagttaat gttgctgctt gcgtccttaa atgacttaag gtttcatctt ccagaagata 1860
ttgagatata ttactgttt 1879
<210> 82
<211> 2767
<212> DNA
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7161479CB1
<400> 82
cccataatat catatgctca atctattctg catttggcct ccgtcatagt cttaatgtca 60
agtacccaaa cactccagga ttcaacaagc cctgaaaact taatagaact ctactttttt 120
agcacttgtg accatttcac cttttcaacc aaagtgactg ttcagtgtta caactttgtc 180
attttgacat gtagcttgca aaggcaatgg attcccatct tggtttcctt aaagtgtaga 240
tgttcaaaaa tatgccagat actctaaatt attggtgtct gttaaatggg aacttctcaa 300
agatcaaaga attgctcaac aagtgctaaa ctttgacata cttcttgcat tttctttcag 360
gctaacaata aatgcccttg cccagaagct caatgcgtac tggaaggaaa aaacatctca 420
agataatttt gagacctcaa ctgtagccag gccaataccg aaggttcctg accagacatg 480
ggttcagtgt gatgagtgtc ttaaatggag aaagcttcct gggaagattg atccatccat 540
gttacctgca agatggtttt gttattataa ttcccatcca aagtacagga gatgctctgt 600
tccagaggaa caagaactca ctgatgaaga cctgtgcttg agcaaagcta agaaacaaga 660
acaaactgtt gaggagaaga agaagatgcc tatggaaaat gagaaccacc aggtattcag 720
taatccacca aagatcctta ctgttcaaga aatggctgga ttgaataaca agacaattgg 780
atatgaggga attcatagcc ctagtgtgct tccttctggt ggagaagaaa gcagatcacc 840
atctcttcaa cttaagcctc tggattccag tgttttacag ttttccagta agtacaaatg 900
gatcctaggt gaagaaccgg tggagaaacg aagaaggctc cagaatgaga tgacaacacc 960
ttctctagat tattccatgc ctgctcctta caggagggta gaagcacctg ttgcctaccc 1020
agaaggggag aacagccatg ataaatcgag ttctgagaga agtacaccac catacctttt 1080
cccagaatac ccagaagcaa gcaagaatac aggtcagaat agggaggttt caattctgta 1140
tccaggggcc aaagaccaac gccaggggtc cctgcttcct gaagaattag aagatcagat 1200
gccaagattg gtggcagaag aatctaacag aggtagcaca accataaaca aagaagaagt 1260
caacaaggga ccttttgtag ctgttgtggg tgttgccaaa ggtgttagag attcaggagc 1320
tcccattcag ctgatccctt ttaacagaga ggagcttgct gagagacgaa aagcagttga 1380
atcctggaac ccagtgcctt attctgtggc ctctgctgca atccctgctg cagccattgg 1440
ggagaaagca agaggctatg aggagagcga aggtcataat acaccaaagt tgaagaacca 1500
gagagagctg gaagaattga agagaaccac agaaaaattg gaacgtgttt tggctgaaag 1560
gaatttgttc cagcaaaagg tggaggagct ggaacaggag aggaatcact ggcagtctga 1620
attcaagaaa gtccaacatg aattggtgat ctacagtacc caggaggcgg aaggcttgta 1680
ctggagcaag aaacacatgg gttatcgcca agctgaattc cagattctga aagctgagct 1740
ggaaagaacc aaagaggaaa agcaagagtt aaaagagaaa ctgaaggaaa cagagacaca 1800
cctggaaatg ctgcagaagg ctcaggtctc ctaccggacc ccagagggag atgacctaga 1860
aagggctttg gcaaagctta cgcggctacg tatccacgtc agctatctcc ttacttctgt 1920
cctccctcac ttggagcttc gtgagatcgg gtatgactca gaacaagtgg atgggatcct 1980
79/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
gtacacggtg ctggaggcaa atcacatact ggattgagca ccagactgta tacccttctc 2040
ttctcttatc ttctgtctgt tctcttttct ctccctccct cacgtctctc tctctctctc 2100
tctctctctc tctctctctc tctcaccctc acctttatgc cttatataga gaatctctgt 2160
gtaaatcctg gctcataatc agtctccttt ttatcagttt tggtgtggag aaagaggcca 2220
gtttaaatag gctttcaaga gtctagggtc agaaaagcaa tagtcactaa gctaggtgac 2280
ctgaaagctt taattttcat gacctggata tgtggtctat tgtatatctt tttctgaaat 2340
ggttgtattc atttaggtta gatcaatcag cagatattgg gtccggtata ccaggtatta 2400
ttttggggta agctaacaag tacaactcat gtttgcagcc ttcgaagatg taacaatctc 2460
gttggaaaca taagacatac atcacattat acacaaaagt gtatgatatg tacaactgag 2520
tggtacagat atgacatgga agatctgggg gaggaagttc aaggaaggca ccacaccaga 2580
aatgggacct aaattaaggc ttaaagaatg agcatggcca cactgtcagt gattgttctt 2640
taggtggtag gactatggtt gatatttttc ttcttcctat cttcctgcct ttttcagatt 2700
ttcaatatta aacttgttat tcttacattg gaaaaataat ggttttaacc gaaaaaaaaa 2760
aaagggg 2767
<210> 83
<211> 2364
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7502313CB1
<400> 83
tcaccacctg ccatacccct taaataagcc cctcaccttg ctgcctcagg accttcaaga 60
ttccatctgt gggctggccg gcaagatggc accagtgggg acccacaccc tggctgggca 120
gaggtgctgc tagcaacctc tcttcctcta taagaggaaa tggaaaatgc agggtgtgga 180
attgcccttt ggggtccttc cttaattgaa ggccaccttc tcacaggttt cattctgcag 240
ggatttactg gaatctattg gtgctgctgc atgagtctgc tgacaacctg actgcacaag 300
gactgggtag cagactcctc agagtcctct tgacacaaat gtcagatttg tgtcactctt 360
ctgccttcgt gaaaagccaa tagcactctc agatatcagg ggattttagt tccaagcagg 420
gaccctggtt tccatactgc cctcagctgg agtttggatc caaaggctct ggctaagtca 480
ttatgtcact ttttcacagg aatgtaaatt tgactgtcac ctctgaattt gttcagtgtc 540
ccaccatggt ctatgagaag tacactggaa gcgtgggggg aacacatgac atgatttgtg 600
aatatcatca tctttgccag acaagtctcc aggggatccc tgtttcccaa ctgaaaggtg 660
tgaacggaca~ cacacacagc ctggatgacg ccttggctgt tctaaggggc tgtaaggtgg 720.
gctctgggcc ttccagctag gctctcaagc acagcagaag cctcactggg ctgctatgtc 780
tctgtatttg tggcttgtgt ggtagcctca gaagcagagc tgtttggcag actggctgga 840
gaaattccct ctaggagact tgcctgtgct gtgcttccag gtcacagagc cccccggaaa 900
ctcacagggg ccctcttccc agaaaagaat ctattctatc acttcagaat caggacactc 960
aagctctggc agaggaaggc caagttactt tcatggtctt accctctgct tttccccttt 1020
ttgcaaaaaa ccactggcca aatccgaacc attgcccttg tttcccccac gttctctctc 1080
agatctttgt ctcgaaggga aaacatagtg gatgaaaagg tgtggcaggc tttggcacct 1140
tgttaaaatt tctagtcatc tgtggatgtt accttgcttg tccacagcag ccagtcaccc 1200
tggccagtcc cacttcctgg ataattctct accctcaccc cacagagcca tctctctcca 1260
gaccaaaagc tggaaggaga gttgctttga gagcttgttt ttacaactgc atgtttatta 1320
tgatactttc tctccaaagg aaacttttaa atcaatggga acaattagca acagaaagag 1380
cacagtccct gcttttgact gggttcctat tttaagcaca aatgagagct ctggagccag 1440
aatgccaggg ttctaacttc agcattcact tactagctgt atgatcttgg ccaagtcact 1500
tcacctccct gagccccaat tcccaagttt gtgaaatggc aacaatacct atgtgtcact 1560
ggattattgg ttaaaacaga atgagattcc ttgtgtgaaa atagctatta tacctgacac 1620
actcatcgta tgggctctgc aaagggatat tccccaacct gtccttcccg acaggaagca 1680
tagggcactg cagatgggga agcatgtcac cttggcagtg actcggtggc ttcccaagca 1740
ggagtgtcag gggaaccatg agagagagtc taggagccaa acacatcacc accctgagca 1800
gatacaggag tggggagggg gctgtaactc agtgagtggc ttccaggggc cccaggccct 1860
gctggatgtg ggccaagccc tacagcttcc ctaggcagta agtaaaaaca ttctcctagc 1920
attaaaatgg tttccataac tacttttgtc ctggcttctt aatactgggt acctggcatg 1980
cagccaagaa ctctgctttt ccgtggtgct tatgtattaa gtagattagc tggggaggga 2040
tattccttgt ttaatggcag atccaggaca ctccggaagc tctgcccacc aacttcacct 2100
taccaggcga gagtagcact gcttggaagg ctgctcctgc cttttaaagc ctgtctacgt 2160
attagctcct ccaccaagga aaagaatttg ctgttagatg gctagggcag gacacggaca 2220
gtcatcaggg gatctatgtt tggcttatgg caagtggctt cactcccacg gctcaggtgc 2280
80/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
cattagggga tattgagccc gcttactaac cccactgacc acctctgcat catttggaaa 2340
atggagagtC tccctgccat tctc 2364
<210> 84
<211> 2597
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7502390CB1
<400> 84
acgctcacac cttgtaatcc tagcactttg ggaggctgag gcgggcggat cacctgaggt 60
caggagttca gcctggccaa catggtgaaa ccccgtctct actaaaaata caaaaattag 120
ccgggcgtgc tggtgcatgc ctgtaatccc atctactcag ggggatgagg caggagaatc 180
gctcaaaccC aaggggcaga ggttgcagtg agccgagatt gcaccactgc actccagcct 240
gggtgacaga gcgagactcc atctaaaaaa aataaataaa catgggagaa acctggaaga 300
atatttgctc aactgtgagg catggttggt ggctaaggga tcacagaatg gcaggcctgc 360
ctattcctcc tgagatcgtt aaggaggctg aggtgccgca ggctgcgctg ggcgtcccag 420
cccaggggac aggggacaat ggccacacgc ctgtggagga ggaggtcggg ggcatcccag 480
taccagcacc ggggctcctg caggtcacgg agaggaggca gcctctgagc agcgtctcct 540
ctctggaggt ccacttcgac ctcctggacc tcactgagct caccgacatg tcggaccagg 600
agctggccga ggtctttgct gactcggacg acgagaacct caacaccgag tccccagcag 660
gtctgcaccc gctgccccgg gccggctacc tgcgctcccc ttcctggacg aggacaaggg 720
ctgagcagag ccacgagaag cagcccctag gcgaccccga gcggcaggcc acagtcctgg 780
acacgtttct cactgtggag aggccccagg aggactagac catctccacc tgccccagct 840
cctgcaggga tggggtccga acacgatggc agatctgggc agtgctgacc cagcagacac 900
acttcacccg cccacgaggc tccagccgtc acctcctgac acacaccctg ggggcagctc 960
tctgccagcc ccgagaccgg ccttgtctgc tgggcacggg tcttcgcctc acttggagac 1020
cagccggctt tcctggggga cacacggggc ccccggatgc ctctgggagc cccagcacaa 1080
gcacagccca gtggccttac gtccagctcg ttcctgggcc ccgagtcagg aagacagcgt 1140
cacggagtca ctgccaggaa cgtgctgagg aatggagtgg cccacggcgg ccttggggtg 1200
aaggggaccc aggcctgtga cagccactcc aggaactcct gggggtgctc caacctccgc 1260
gttttcctgt gctgccaagc tcagaagcca gaggcgggtt tggtagtggc taatgggaca 1320
atgtgctgtc cagcaaagca Cacatggaga agcggcccca aaattcccac cttgatttcc 1380
atcctgcccc ttcttctact ccacggaggc gctgtctcac tagggtcccc tccccaaggc 1440
tcagctctaa gacctgcacc tgcttctctt ggcccctgcg tgacagacaa gtccattccc 1500
tccttagctc agaacaccaa atatcaccag actgcctaag agacttgatg acacctcccg 1560
gaatgctctc ggggtggggt tcacctctcc ttgtcctgca cccactgcta ggccacattc 1620
tcgtttctgc tcacatccca ttgcccggct acaaggcctg cccacggccc ttaaacttgc 1680
tgggcaggtt tggagcccat gggaccccgt gggtctctgt ccaggagcag cagaggaggc 1740
tgacaggccc tgctccctct gctctggggg tgtctgggag ccccagctca caccctccca 1800
atgcttatat gctgaagctc acagaatggg cttcttgcct gacagcaagt caaagaatga 1860
gtttaatatc aaagtgtaag cttactttcc atccccaagc cagcctgccc cctgccccat 1920
ttcccatgag cacacttctg gggaaggaaa acaggctcct ggccttcact ctcagcagag 1980
ctttggagat gccccaggca tgccctgagc tccttctgtg tacctgctcc cacttctgag 2040
ccacccgctg cccctccgca ctgctggcaa acccagttcc tgcctcagcc aggtctcctt 2100
ccctggtttc cagtcacaca gagcccagca gctttctctt tcagtcccat aagggcagcc 2160
ttgtgtccct ggccacactt ccacccgcca gggtcttcct ccccatcttt ccatccttcc 2220
tgctgagctt ccacagagct cgtttgcaaa cagggggatt aaagcatcac tgcgcattcg 2280
aaggcctggc cacagtctct tcctttccat agcagagtga acgaggtgcc tgctgagggg 2340
ttgtgagctg agctgcctgg gctccgtgcc cggaccattc tccagctgca gcagcctgag 2400
ggctctgctg tgctcacttg ggtcacatgt gggggatgag tgacctggat ttccagtccc 2460
aaagtcacac acagaaggaa tcattaaaga agtgaaagtc ccgccccacg tgcacgtaaa 2520
ggtagcccag gctgggggag ggagtcacag aagtcccacc ccacgtgcat gtaaaggtag 2580
cccaggctgg gggaggg 2597
<210> 85
<211> 2229
<212> DNA
<213> Homo sapiens
81/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<220>
<221> misc_feature
<223> Incyte ID No: 7502872CB1
<400> 85
ggccgggctg ggatagcgcg agtgtccgcg gccgagcagc agagattttt gctgtgagaa 60
ttaattacca gtaacagttc aatatggggg acattctggc tcatgaatct gaattacttg 120
gactagtgaa agagtattta gattttgctg aatttgaaga caccttgaaa acattttcaa 180
aagaatgcaa aataaaagga aaaccattgt gtaaaacagt aggcggatct ttcagagact 240
ccaaatcatt gacaattcag aaggatcttg tcgctgcatt tgacaacgga gaccagaagg 300
tgttcttcga tctgtgggag gagcacattt caagttccat ccgagatggg gactcctttg 360
cccagaagct ggaattctat ctccacatcc attttgccat ctatcttttg aagtactctg 420
tggggagacc ggacaaagag gagctggatg aaaagatttc ctacttcaaa acctacctgg 480
agaccaaagg ggcagccttg agccagacca cagagtttct tcctttctat gcccttcctt 540
ttgttcccaa ccctatggtg cacccctcat ttaaagaact cttccaggat tcctggactc 600
cagagttaaa gttgaagttg gaaaagtttc tagctttaat atctaaagcc agcaacacgc 660
caaagctttt aacaatatat aaggagaatg gacaaagtaa caaagaaatc ttgcagcagc 720
tccaccagca gctggttgaa gctgaacgta ggtcagtgac atacctcaaa cggtacaata 780
agatccaggc cgactaccac aatctcattg gagtcacagc agagctggtg gattctctag 840
aggccacagt cagcggcaag atgatcaccc ctgagtacct ccagagcgtc tgtgtccgcc 900
tgttcagtaa ccagatgcgg cagagcctgg cgcatagtgt ggacttcacg aggcctggga 960
cggcatccac catgttacga gcctccttgg cacccgtgaa attgaaggat gtcccattac 1020
tgccctcctt ggattatgag aaactgaaga aggatttgat tttggggagt gaccgcttga 1080
aagccttctt gttgcaggct ctgcgctggc gcttgaccac atcccatcct ggagagcaga 1140
gggagaccgt tctgcaagcc tacatcagca atgacctctt ggactgttat agccacaacc 1200
agaggagtgt gcttcagttg ctgcactcca cgagcgacgt ggtgcggcag tacatggcca 1260
ggctcatcaa tgcttttgcg tcactggcag aaggtcgcct ctaccttgcc cagaacacaa 1320
aggtgctgca gatgctggag ggaaggctga aggaggagga caaggatatc atcaccaggg 1380
agaatgttct tggggccctg cagaagttca gtctcaggcg cccgctgcag acagcgatga 1440
ttcaagacgg cctcatcttc tggctggttg atgttctgaa ggaccctgac tgcctgtctg 1500
actacacgct ggagtactcg gtggctttgc tcatgaacct ctgcctccgc agcacaggga 1560
agaacatgtg tgccaaggtg gcaggcctcg tgctcaaagt cctttcggat cttcttggcc 1620
atgaaaacca tgagatacag ccgtatgtga atggagctct gtacagcatc ctttctgttc 1680
catccattcg tgaggaagca agagcaatgg gaatggaaga catcctacgc tgcttcatca 1740
aagaaggcaa tgctgaaatg atccgccaga tagaattcat catcaagcag ctaaattccg 1800
aagagctacc agatggtgtt cttgaatctg atgatgatga agatgaagat gatgaagagg 1860
accatgacat catggaagcc gatctggaca aagacgaact gatccagccc cagctcggag 1920
aactctcagg agagaagctt ctgaccacgg agtacctggg gatcatgacc aacacgggga 1980
agacaaggcg gaaggggctg gctaatgtgc agtggagcgg ggatgagccc ctgcaaaggc 2040
ccgtcacccc cggcggccac agaaacgggt acccagtgta agtcagggct aaaggaagcg 2100
ggaattgact ttcttaagct ttgttttgat tacagtgtaa gatgtatgta tttttaaaat 2160
tcaaaataaa gcattcattt tgaaacaaaa aacaaaaaaa aacacacaca acaccacaac 2220
aacacaaaa 2229
<210> 86
<211> 2504
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7505443CB1
<400> 86
gtttgaaaca tggcgcgggc tggccctcgg ctggtgctga gcgaggaggc ggttcgggcg 60
aagagcggct tagggcctca ccgcgacctg gctgagcttc agtcattgtc tattcctgga 120
acttaccaag agaagatcac ccacctggga cattctctga tgagtttaac aggtctgaaa 180
tctttggatc tctcgcgcaa ctccttggtt agtctggagg gcattcagta cctgactgca 240
ttggagagtc tcaatctcta ctacaactgc atctcctcgt tggcagaagt gtttcggctc 300
cacgccttaa ccgagctcgt ggatgtggac ttccggctga accccgtggt gaaggttgag 360
cctgactacc gcctttttgt tgtgcacctg ctccccaagc tccagcagct ggaatccaga 420
catctgttga gcccgcagtt ggtacagtac cagtgtgggg actctgggaa gcagggccgt 480
gagacgagga ggagcagctg cagagggtgc tgtctggaga agatgccttg gagccagctc 540
82/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
tgtggagagc ttccgccact gtacggagcg gagccagagg cctcccgtgc ccccaggcca 600
cacacgtact tcaccccaca cccagactcc atggataccg aggactcggc ctcttctcag 660
aagttggatt tgtcaggaga aatggtgcct ggtcccctgc cagcccccgg aaagtgcagg 720
aagcgaagaa tgcctgttgg aagattccag acgttttcgg accaggaggg tttgggctgc 780
ccggagagaa ctcatgggtc ctccgtgccc aaggagagcc tgagcagaca ggacagctca 840
gaaagcagga acgggaggac cttgtctcag cctgaggcct cggagactga ggagcagagg 900
tctcggggtg tgaccgacac cagagagccg tctcccgggt cacactcggc tctacccggg 960
aagaagacgg ccctgcaggc ggcgctcctg gagacgctct tggacctggt ggacaggagc 1020
tggggcggct gcaggtccct gcacagcaac gaggcattcc ttgctcaggc aagacacatc 1080
ttgtcatctg ttgaagaatt cacagcagct caggacagct ctgcgatggt gggtgaagat 1140
gtcggctccc tggctctgga gagtaagtcc ctgcaaagcc gccttgctga gcagcagcag 1200
cagcacgccc gggagatgag cgaggtgacg gcggagctgc accacgcaca caaggagctg 1260
gatgatttga gacaacattt agataaatct ttggaagaga acagtaggtt aaaatcgctt 1320
tggttgagta tgaaaaagga agtgaagagt gcagacactg cagccacgtt aaatttgcag 1380
atcgctggac ttcaaacaag tgtgaagagg ctgtgtggcg agattgtgga actgaagcag 1440
cacctggagc actacgacaa gatccaggag ctcacgcaga tgctgcagga gagccacagc 1500
tccctggtca gcaccaatga acacctgctg caggagctga gccaggtgcg ggcgcagcac 1560
agagccgagg tggagcagat gcactggagc taccaggagc tcaagaagac catggccctg 1620
tttccacaca gcagcgccag ccatggaggc tgccaggcct gctgactcct gccgagaagc 1680
tgggccaccc cttaagcttc ctggtaaagt tacattgtct gcacctttgt acttctttac 1740
tgagtgtact ggctggcaag agttctctct tctgttggta attatttagg atttttggaa 1800
tgtattcagg acctgtagct tggttttcta aagcacctcg taaaatgata tgattactcc 1860
aagcccctct gcatgttttc agacagaaca cattgacata ttttgagaca aactgactat 1920
taatcttgta tccagtatcc tgagatgaag taaatgcagt gttctactgc ctgatgtgaa 1980
agagagctat gtatgataat taaagaaaat aattttctgt gtaacaagca atctttattt 2040
aataaacaaa taacattgtt ctgaaaagtt aactttttca gtggctgtat acaaattata 2100
actgagtttg tcattgagtt ttttatagaa caagctgtgc accatgatag gtgagtggga 2160
ataattaagt tcaaagactt aacacaagtg acacttatag atgtggggga aaaaacctta 2220
aaaatattgt cttatgatat taacatatca cagcggaaac atctacccaa gcctggctcg 2280.
ctctcttgtt tgcagtgtag ttctgaaaat tatgtcataa taggacacaa aactagattt 2340
ttagtttgga aacttctata tcttatgtct atacttgctg tgattggtaa gacaaggtca 2400
ttttttaaat gacgtacaac taatttggga agaatgagca ggcctaagaa gaaaatgctc 2460
tgtgctagct ctgtgtgtgt gtgtgccctt gctgtgtgtg tgcc 2504
<210> 87
<211> 701
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 8032443CB1
<400> 87
ggggaccatc tagcccctaa ccatgggccc agaagagaag accatcatga cagaaaggtc 60
tgcagctgtt ttcatccagg cctggtggcg gggcatgctg gtgcgacgca cactgctgca 120
tgcagccctc agggcttgga tcattcagtg ctggtggagg caggtgctgg agaagctgct 180
ggcaaagagg cggaggatgg tgttggagtt ctatgtgcag caggaatggg cagcagtcag 240
gctgcagtcc tgggtccgca tgtggtgtgt ccgccagcgt tactgtcgtt tgctcaacgc 300
tgtccgcatc atccaggtct attggcgctg gcacagctgc cattcccgtg tctttattga 360
gggccactat gaactcaaag aaaaccaact taatattcaa cttgaaatct ctttgggctt 420
acaggcttgt aaggtgcaac aatgcatacc ccttccatta aaagaatgac caggtctgct 480
aaaaaaaaaa aaaataacag cagccatcag aagacagcag caaaactaag caaaaacagg 540
ggggggcggg cttacttaga gagctccagg tagttgttaa gcatttgtgg agacatcgcc 600
ctctattagg tggcccaaaa aatagacatg aggggggtgt tttcgcgccg tgcggtggaa 660
ccccggtgtc cccttttccg cgtggctccc tcctgcgaca a 701
<210> 88
<211> 1569
<212> DNA
<213> Homo Sapiens
<220>
83/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<221> misc_feature
<223> Incyte ID No: 7704916CB1
<400> 88
ttaacacatg tcaagtacct agaacagtcc cttttattcc atcacatggg ttgcgaggtg 60
aagtgcacca ttgccttgac cctggggaac ttgataacca aatacagaat taaattgctt 120
accaaacccc atttatattt atagctggaa gagcctgtat tgtcctcaca atagtataga 180
agaattcaag agaggagaga gagacagcac cgaatgaaga ctgtaaaaga aaagaaggaa 240
tgccagagat tgagaaaatc tgccaagact aggagggtaa cccagaggaa accgtcttca 300
gggcctgttt gctggctatg ccttcgagaa cctggggatc ccgaaaaatt aggggaattt 360
cttcagaaag acaatatcag cgtgcattat ttctgtctta tcttatctag taagctgcct 420
cagaggggcc agtccaacag aggtttccat ggatttctgc ctgaagacat caaaaaggag 480
gcagcccggg cttctaggaa gatctgcttt gtgtgcaaga aaaagggagc tgctatcaac 540
tgccagaagg atcagtgcct cagaaacttc catctgcctt gtggccaaga aaggggttgc 600
ctttcacaat tttttggaga gtacaaatca ttttgtgaca aacatcgccc aacacagaac 660
atccaacatg ggcatgtggg ggaggaaagc tgcatcttat gttgtgaaga cttatcccaa 720
cagagtgttg agaacatcca gagcccgtgt tgtagtcaag Ccatctacca ccgcaagtgc 780
atacagaaat atgcccacac atcagcaaag catttcttca aatgtccaca gtgtaacaat 840
cgaaaagagt ttcctcaaga aatgctgaga atgggaattc atattccaga caggaggtgg 900
tgcctcattc tgtgtgctac atgcggatcc cacggaaccc acagggactg ctcctctctt 960
agatctaaca gtaagaaatg ggagtgtgag gagtgttcac ctgctgcagc cacagactac 1020
atacctgaaa actcagggga catcccttgc tgcagcagca ccttccaccc tgaggaacat 1080
ttctgcagag acaacacctt ggaagagaat ccgggccttt cttggactga ttggccagaa 1140
ccttccttat tagaaaagcc agagtcctct cgtggcagga ggagctactc ctggaggtcc 1200
aagggtgtca gaatcactaa cagctgcaaa aaatccaagt aacaccttct gagtagctgc 1260
tgtcccacac aatagggtat gaagctgcgc tcctccatcg ggtttgggga gggagcactc 1320
tgggactgtg agacaaggaa gcagggccag cagtgagact atgagccaag caaagagaag 1380
tctcagtgga gcatgaggag ggagcagtcc agatgccaac aaggaaatgc gtttatggct 1440
acaagagtgc ctctgctttc tcctcctctc ctcccaccaa ggattcttcc accttaatct 1500
tgttttcata tgcctcttct tacttcaccc atgtttgttg ttatgcaaat aaaggttttc 1560
tctccaaaa 1569
<210> 89
<211> 1052
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 2013440CB1
<400> 89
ggtagagaat ccaagataga tcaactctcc ctaaaggctg acagtgaact cttggggccg 60
ttttattctc tgaggttagc aaggagtcat ctactagcca ttcaggaggc cagctgggaa 120
gacaaaatag gcaccccaaa ctcagcaact tcataacacc ttcctctccc cgcctgaagc 180
cttaaactgc atcaagtcaa agaaacctgg ggcaaatcct taacatgttt ttgactgcag 240
taaatccaca gccactctct actccgagct ggcagattga gaccaagtat tcaacgaaag 300
tgctcactgg aaattggatg gaagagagga gaaagttcac cagagacact gacaagacac 360
cccaatccat ttacagaaaa gaatacatcc ccttcccaga ccacagacca gaccagatct 420
ccaggtggta tgggaagagg aaagttgagg ggctacctta caaacacctg atcacccacc 480
accaggagcc cccacatcgc tacctgatca gcacctatga cgaccattac aaccggcatg 540
gttacaaccc ggggctgcct ccactccgca cttggaatgg acagaagttg ctgtggctgc 600
cagagaagtc tgactttccc cttcttgctc cccctacaaa ctatggactc tatgagcagc 660
tcaagcagag acagctcaca cccaaggctg gcctgaagca gagcacttat acttcatcct 720
accccagacc accgttgtgc gctatgtcct ggagggagca tgcggtcccg gtccctcccc 780
atcgcctgca tcctctccca cacttctgag agctgccacc ccaggagcag ctcagataga 840
atcagctgga gaccacagca tcactggact tgccagacaa caagtggcgc agataaactc 900
agagtacgag atctggcccg tcaaaggtgc tctcagaatc atcatctgca tttggcggta 960
cctgtccccc ctcaaaaccc acaggttcct ttcttttcca tccaacaatt aaagatcttt 1020
gacactaaaa aaaaaaaaaa aaaaaaaaat tg 1052
<210> 90
<211> 1325
X4/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 2503512CB1
<400> 90
ccctgaccct cagcagccac aggcctccgc cgaagcccca tttgctgcca gagggatcta 60
ctcggaggag atgccgtcgg tggcccggcc tcggcctgtc gggggtacca caggctccca 120
gatccagcac ctgacacagg tggggattgc cagcagaatt ggagctcagc cagtggaaat 180
cccgccaagc agaggcagcc agtatggggg gccaggctgg ccttcgtacg gggaggacga 240
agcggggcga agagaggcca cacacatgct cggacatcaa gagtattctt cttcaccgct 300
atttcaggtg ccaaggactt caggcaggga gccctcagct ccttccggga acctccccca 360
ccggggactg cagggccctg ggctgggtta ccccaccagc tccacggaag acctccagcc 420
tggccactcc tcggcctctc tcatcaaagc aatccgcgag gagctcctcc ggctctccca 480
gaaacagagc accgtgcaga acttccacag ctgatcggcc tcgcctcgca gatttgccaa 540
gtatccgctt cctgtggaag caagaccaaa aggaaatcaa ctgagtgggt gtttggaaga 600
ggaaggagca actctcgggc agcctgccca agggagggag caagttgcaa tttagaagat 660
gccatacgtc gtgtgacagc tcatgagcct ttcactgggc tggcaattgt ctgaacactt 720
gggttcagtt gaaatatatg tattttggcc aaaagccagc agcacttcac aaaaacaaaa 780
cacaaaccta agctaacaaa atgactgcat tcgtctcttt tttaaaggta gagattaaac 840
tgtatagaca gcatagggat gaaaggaacc aagcgtttct gtgggattga gactggtacg 900
tgtacgatga acctgctgct ttgttttctg agaagaggtt tgaagacatt ttattaacag 960
cttaattttt ctcttttact ccataggaac ttattttaat agtaacatta acaacaagaa 1020
tactaagact gtttgggaat tttaaaaagc tactagtgag aaaccaaatg ataggttgta 1080
gagcctgatg actccaaaca aagccatcac ccgcattctt cctccttctt ctggtgctac 1140
agctccaagg gcccttcacc ttcatgtctg aaatggaact ttggcttttt cagtggaaga 1200
atatgttgaa ggtttcattt tgttctagaa aaaaaaaatc cctcccaaag tggggcaaaa 1260
agctttatat ttatttgatt atccaaaata cagatcaaag tttagatcta caaaaaaaaa 1320
aaaaa 1325
<210> 91
<211> 2110
<212> DNA
<213> Homo Sapiens
<220>
<221> ~misc_feature
<223> Incyte ID No: 277396CB1
<400> 91
caagaattcg gcaccgggga ctgcggaggc cggggcggtg gcgtgcaggg tgaggtctcc 60
cgggctggcg cgcgtgcaag ccccctgctg ctacctcgcg ctaggccagc tccaggaggg 120
caacagacga caggacattt caaaagggcc acttaaattt tccttcgagt aaacagaaca 180
cctgccaaga ccctcgagcc tgggcgtgcc ccaagacagg tggttggggt ctaggagtcg 240
gctgcagctg gcgatgctct gaggggacgt cagggcctca ggatcgcccc tatgaagacg 300
aagtgcatct gtgaactgtg ctcctgcggg cggcatcact gtccacatct ccctaccagg 360
atttatgatg aaacagagaa accatgtctt ctctccgaat ataccgagaa ctaccctttc 420
tatcactcct acctgcccag agagtccttc aagccaaggc gggagtacca gaaagggtct 480
ataccaatgg aaggcctgac tacatcaagg agagattttg ggcctcacaa agtggcacca 540
gtgaaggtcc accagtatga ccagttcgtc ccgagtgaag agaatatgga tttgctcacg 600
acgtataaga aagattacaa tccctaccct gtctgtcgag tggaccccat caaacctcgg 660
gacagtaaat atccatgtag cgacaagatg gagtgtttgc ctacttataa agctgattat 720
ttgccttgga accaaccaag gcgagagcca cttcgtctgg aacacaaata ccagccggca 780
tcagtcaggt ttgataacag aaccacacac caggacgatt accccataaa aggccttgtg 840
aagaccataa gctgtaaacc tctggccatg ccaaagctct gtaacatccc cttggaggat 900
gtgactaact acaagatgag ctatgtggcc caccccgtgg agaagcgctt tgtgcatgaa 960
gcagagaagt tcaggccctg tgaaatcccc tttgaaagcc ttaccactca aaaacaatcc 1020
taccggggcc tgatggggga gcctgccaag agcttgaaac ctctagccag gcctcctggg 1080
ctagacatgc ctttctgtaa caccactgag tttcgagata agtaccaagc ttggccaatg 1140
ccccggatgt tctccaaagc tcccatcacc tacgtccctc ccgaagacag gatggatctt 1200
ctgacaacag tgcaggccca ttacacatgc cctaagggtg ccccagctca gtcctgccga 1260
X5/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
cctgcacttc agattaagaa gtgcggtcgc tttgaaggct cttccaccac caaggatgac 1320
tacaagcagt ggtccagcat gcgcacagag ccagtcaagc ccgttcccca gctggacttg 1380
cccaccgagc ccctggactg cctgaccacc actcgggccc actatgtgcc ccacctgcct 1440
atcaatacca aaagctgtaa gcctcattgg tctggccctc gaggaaatgt ccctgtggaa 1500
agccagacca cctacaccat oagctttact cccaaggaaa tgggcaggtg cctagcttca 1560
tatcctgagc ctcctggcta cacctttgag gaagtggatg ctttgggtca caggatatac 1620
aaaccagttt cccaggcagg ctctcagcag agcagccatc tttctgtaga tgattcagaa 1680
aaccccaacc agagggagtt ggaagtgtta gcctgatttt gaaaaatagt aatttagaaa 1740
ttacacagta cttttaaaag cagacaactg agaattattt gttggacaaa aaagaattcc 1800
ctaaaatgac aaaaaacaaa aaacaaaaaa ccttcaccac ttccagagca cttgaataaa 1860
atgagaatca cttgactcag ggaaaatgac atttaatcac cggctaatta actcccctta 1920
CCttCtCCtt taCtgCatCC CCdCCCCgtt tCaggtCCtt taCtttgtgC ttatggaatc 1980
aaagttggtc tgtgaggttt tctctgatcc agatgtattt tattaattta atcattgtat 2040
gaactgacat aatgattaac ccttgccaag tatgaagcgc caaagaataa attttatttt 2100
ggggggaatg 2110
<210> 92
<211> 1927
<212> DNA
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 3044046CB1
<400> 92
cggagccaga gagacgcagc taggcctcca cggctgtgga gagatcctgc cacgggcctt 60
gttcaccatg tcggtgctgg atgcgctttg ggaggatcgg gatgtccgtt tcgacctgtc 120
cgcgcagcaa atgaaaacaa gacctggaga agtccttatt gattgtttag attccattga 180
ggacaccaaa ggaaataatg gagatagagg tagactcttg gtaacaaatt taagaattct 240
ctggcactct ttggcattat caagagtcaa tgtttctgtc ggttacaatt gcatattgaa 300
tattacaaca aggactgcta actctaaatt acgaggccaa actgaagctc tctatatact 360
aacaaaatgt aacagtactc gttttgaatt tatatttaca aatttggttc ctggaagccc 420
tagacttttt acttctgtga tggcagtaca cagagcttat gaaacttcta aaatgtatcg 480
tgattttaaa ttaagaagtg cactaattca gaacaagcaa ctaagactgt tgccacaaga 540
acatgtatat gataaaataa atggagtttg gaatttatcc agtgatcagg gcaatttagg 600
aacctttttt attaccaatg tgagaattgt gtggcatgca aatatgaatg atagttttaa 660'
tgtcagtata ccatatctgc aaattcgttc aataaagatt agagattcaa aatttggttt 720
agctcttgtc atagaaagct ctcagcagag tggtggatat gttcttggct ttaaaataga 780
tcctgtggaa aaactacaag aatcagttaa ggaaatcaat tcacttcaca aagtctattc 840
tgccagtccc atatttggag ttgattatga gatggaagaa aagccccagc cgctcgaagc 900
tctgacagtc gaacaaattc aagatgatgt agaaatagac tctgatggtc acacggatgc 960
ttttgtggct tattttgctg atggcaataa gcaacaagat cgtgaacctg tattttcaga 1020
agaactgggg cttgcaatag agaaattgaa ggatggattc accctacagg gactttggga 1080
agtaatgagt tgattgacct tgagttgaga tggatttcta ttaaagatat ctctagttta 1140
aagatactag tcacctgcca taagtcatgg aatagttttt atatttacag cttttatatt 1200
taaaacttgt aagagttttt ttaatgattg aggaaaaagt catttagaaa acttcagttt 1260
tcggccagcg cgtcgaggga ggggccagcg acacatggcc tagtaaccgt ccggccgcgg 1320
cgctggctta agccatggct gagggtagct ggattcctca ggcccgggcg ctcctacagc 1380
agtgcctgca cgcccggctg caaattcgcc cagccgatgg ggacgtcgcg gcccagtggg 1440
tggaggtcca aagaggactg gtgatctacg tgtgcttttt caagggagct gataaagaac 1500
ttcttcccaa aatggccgaa gctggactgt actgctgcca tctctggctc actgcaacct 1560
ccctgcctga ttctcctgcc tcagcctgcc gagtgcctgc gattacaggc gcgcgccgcc 1620
acacctgact ggttttcgta tttttttggt ggagacgggg tttcgctgtg ttggccgggc 1680
tggtctccag ctcctaaccg cgagtgatcc gccagcctcg gcctcccgag gtgccgggat 1740
tgcagacgga gtctggttca cttagtgctc aatggtgccc aggctggagt gcagtggcgt 1800
gatctccgct cgctacaacc tccacctccc agctgcctgc cttggcctcc cgaagtgcca 1860
agagtgcagc ctctgcccgg ccgccacccc gtctgggaag tgaggagcgt ctctgcctgg 1920
1927
ctgccca
<210> 93
<211> 1051
<212> DNA
86/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 3808420CB1
<400> 93
ggagtcgcgc aggcaggcgg aggctgaggg cgccgctggc cggccctccc agccctctcc 60
gcgcggctcc gccggggttc caagaggagc tagtaggttc ggtgggggcc ctggcatgga 120
catgcacagc gctcggcttg acagcttcct tagccagctc cgctgggaac tgttgtgtgg 180
tcgggacaca ggctcaccct caatgcctgg tcccctgcag ccaacctccc aaactggccc 240
agatgtgcag cccagccacc agcttagggc ctcgggtgcc ttggaagagg actcagtctg 300
ctgtgtggag gaggaggaag aggaggagga ggaagcagtg gtgacagaag acagggatgc 360
agccttggga ggccccaggg agcatgccct ggactgggac tctggcttct cggaggtgtc 420
aggcagcaca tggcgagagg aagaactgcc tgtatcccag cgcccagcac cctcagcaca 480
gccccttcgt aggcagtgcc tctcagtcag tggcctcccc atgcccagca gggcccctgt 540
agccagtgta ccacctgtcc accatccacg gcccaagtcc accccagacg cctgcctgga 600
gcactggcag ggactggaag cagaggactg gacagcagcc ctactgaaca ggggtcgcag 660
tcgccagccc ctggtactag gggacaattg ctttgctgac ttggtgcaca actggatgga 720
gctgcctgag acagggagtg aagggggtga cggaggtggg caccgtgccc gtgctcggcc 780
ccctcagttc ctgcttggcc tctctgagca gcttcggcgc cggctggcca gggctcggcg 840
gacagctatg gcaggaaagc ggctgtcatg cccacctcgc ccagaacctg aactgcctgc 900
ggatgtctca cgctttgcag ctctcatgag ctgtcgtagc cgccagccca tcatctgcaa 960
tgatgtcagc tacctctgac cctgccctcc agcctgggac aataaaagcc ttttttctag 1020
acaaaaaaaa aaaaaaaaaa aaaaagggcg t 1051
<210> 94
<211> 2328
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7504028CB1
<400> 94
ggctcggaaa tctagttcgg gaaaagtgtg aggggctgtt cacgtgggga aggaacagca 60
ggcgcggagg agggggcaag cgtgtgtgag attcagtggt ccatgcgtgc gtttgtcgtg 120
taagggtcat tcctggggtt tggagtgggg gaacaaatca atgtggctgt ttttccgtgg 180
aaagaattcc cactgcagtg tcccggagcc tgcgtgtggt gggcaagctc ctcagatggt 240
atctcacagg gaatagggga gtcttgaaaa cgcagcttcg gcagtaggaa catgaacctc 300
ttacctaaaa gttccaggga gtttggctcc gttgactatt gggagaagtt cttccagcag 360
cgaggaaaga aagctttcga gtggtatgga acctacctgg aactgtgcgg ggtgctacac 420
aaatatatca agcccaggga aaaggtgctg gtgattgggt gtggcaactc agaactgagt 480
gagcaactgt atgatgtggg ctatcgggat atagtgaaca tcgacatcag tgaggttgtc 540
atcaagcaaa tgaaggaatg taatgccacc cgacggcccc agatgagctt cttgaagatg 600
gacgtgacgc agatggagtt tcctgatgcc tcgttccagg tggtgttgga caagggcacc 660
ctggatgctg tcctgacaga tgaggaagag aagaccttac aacaggtgga caggatgctg 720
gctgaggttg gccgtgtcct gcaggtgggc ggtcgctatc tctgcatctc cctggctcag 780
gctcacatcc tgaagaaagc agtgggccac ttctcccggg aggggtggat ggtgagggtg 840
caccaagtgg ccaacagcca ggaccaggtg ttggaagcag agcctcagtt ctccttgcct 900
gtctttgcct tcatcatgac caagttcagg ccagtccctg gctctgccct tcagatcttt 960
gagctgtgtg ctcaggagca gcgcaagcct gtgcggctgg agagtgccga gcggctggcc 1020
gaggcggtgc aggagcgaca gcagtatgcc tggctgtgca gccagctgcg ccgcaaggcc 1080
aggctgggga gtgtgtctct ggacttgtgc gatggggaca cgggggagcc acgctacacc 1140
ctccacgtgg tggacagccc cactgtgaaa ccatcgcggg acaatcattt tgcgattttc 1200
atcatccctc agggccggga gaccgagtgg ctctttggca tggatgaggg ccggaaacag 1260
ctggcggcca gtgctggctt caggaggttg attacagtgg cccttcaccg aggtcagcag 1320
tatgaaagca tggaccacat ccaagctgag ctgtcggcta gagtcatgga gctggcccca 1380
gctgggatgc ccacccagca gcaggtcccc tttctgtctg tgggtgggga cattggggtc 1440
cggaccgttc agcaccaaga ctgcagcccc ttgagcggtg actatgtcat tgaggatgtg 1500
caaggggatg acaagcgata cttccgtcga Ctgatcttcc tcagcaacag gaatgtggtg 1560
cagtccgaag ccaggttgct gaaggatgtg tctcacaaag agatcccact ggcattgttg 1620
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
gtggtaggcc tgggcggggg cagcctcccc ctctttgtcc acgatcattt tccaaagtcc 1680
tgcattgatg ctgtggagat cgatccctcc atgttggaag tggccaccca gtggtttggc 1740
ttctcccaga gtgaccgaat gaaggtccac attgcagatg gcctggacta tatcgccagc 1800
ttggcaggag gaggagaagc acggccttgc tacgatgtca taatgtttga tgttgacagt 1860
aaggacccaa cactgggaat gagttgtccg cccccagcat ttgtggagca atcttttcta 1920
cagaaggtta aaagcatctt gactcctgaa ggtgttttta ttctcaacct tgtgtgccga 1980
gacttggggc taaaagactc agtgctggct gggctcaagg cagtgttccc cctcctatat 2040
gtccggcgaa ttgagggtga agtgaatgag atcctgttct gtcagctgca ccctgagcaa 2100
aaacttgcca caccagagct cctagaaaca gcccaggctt tggagcggac cctgaggaag 2160
cctgggaggg gttgggatga cacgtatgtc ttgtcagata tgctcaagac ggtgaaaatt 2220
gtgtgactgc ttaggccaag cagccctcct gcctagactg accttggact cccagcctgc 2280
cagagaatga agaaatacaa cgcacagtaa aaaaaaaaaa aaaggggg 2328
<210> 95
<211> 4782
<212> DNA
<213> Homo Sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 7766880CB1
<400> 95
caagaatcaa cttataaggt gtcaaaggca gatgacagat attctcagag tgtaatcaga 60
agtaattccc gtcttgaaga tcaagttatt ggggttgctc tgcaagcatc aaaaaaagaa 120
gaaagtgttg ttggttcagt gacacaactt aaccaacaaa ttggccaagt caataatgca 180
gctacccttg atcttaagaa ctcaactaat ttaatacaga ctccacaaat aaggttgaat 240
actaaagact taaagcagca acatcctctc atacttaagg tgcatgagtc caaggtccag 300
gaacagcacg atcaaataat taatgcttca tctcagattc aaattccaaa tcatgcttta 360
gggcatggcc atcaggcatc tcttcctaat acacaggtcc ttttagattc tgcctgtgat 420
ttacaaattc ttcagcagtc aatactgcag gcaggtttag gtcaagtaaa ggcatcttta 480
Caagcacagc gtgttcaaag ccctcaacaa atagtacatc ccttccttca gatggaaggt 540
catgttattc aaagcaatgg tgatcattct cagcagcaac tccatcctca aaattctgaa 600
gttatgaaaa tggacctctc tgagtcttca aaaccattac aacaacatct aacaacaaag 660
ggccatttta gtgaaacaaa tcaacatgat tcaaagaatc agttgtttct cttggatcga 720.
tgtgtttccc agaggcaggc ttcttagtga tgaaagaaat attttatcaa atgtagatga 780
tatcttagca gctacagcag cagcttgtgg agttacacct actgattttt ccaagtcaac 840
ttcaaatgaa accatgcagg ctgttgaaga tggtgattct aaatctcatt ttcagcagtc 900.
attagatgtc aggcatgtga cttcagattt taactctatg acagctacag taggaaagcc 960
acagaatata aatgatactt ccttaaatgg aaatcaggtt actgtgaacc tttcaccagt 1020
acctgccctt cagtcaaaaa tgactcttga tcaacagcac attgaaacac ctggtcaaaa 1080
tataccaact aaagtaactt cagcagtggt tggaccaagt catgaagtcc aggagcaaag 1140
ttctggccca ttcaagaaac agtctgctac caatcttgaa tctgaagaag acagtgaagc 1200
tcctgttgat agtacattaa ataataacag aaaccaagag tttgtttcta gtagtagaag 1260
tataagtgga gagagtgcta catcagagag tgaatttacc ttagggggtg acgacagtgg 1320
tgtgtcaatg aacccagcta ggagtgcact tgcactgttg gccatggccc aatctgggga 1380
tgcagtcagt gtcaagattg aagaagaaaa ccaagattta atgcatttta accttcaaaa 1440
gaaaagagct aaaggaaaag ggcaagttaa agaggaagac aacagtaatc agaaacagct 1500
gaaaagacct gcccaaggca aacgccagaa tccaagggga acagatattt acttaccgta 1560
tactcctcct tcctcagaaa gctgccatga tggttatcag catcaagaaa aaatgagaca 1620
gaagatcaaa gaggtggagg aaaaacaacc ggaagtcaaa acaggattta ttgcttcttt 1680
cttagatttt ctgaaatccg ggcccaagca gcagttttcc actcttgctg tacgaatgcc 1740
taacaggact agacggccag ggacccagat ggttcgtaca ttttgtcccc caccacttcc 1800
caagccttca tctacaacac ccacaccttt agtgtctgaa actggcggta acagtccatc 2860
agataaagtt gataatgaac ttaaaaactt ggaacattta tcttcatttt cttctgatga 1920
agatgatcct ggatatagtc aagatgctta taaaagcgtc tctactccct taactacttt 1980
ggatgctact tctgataaaa agaagaaaac agaagcccta caggtggcaa ctactagccc 2040
aactgccaat actactggta ctgctactac ttcctcaacc actgtgggtg cagttaagca 2100
agaacctctc cactctactt catatgcagt aaatattctg gaaaatataa gctcttcaga 2160
atcctcaaag cccattgaac ttgatggtct tccttcagac cagtttgcaa aaggacagga 2220
cactgttgcc atagaaggtt ttacagatga ggaggacaca gaaagcggag gagaaggcca 2280
atacagagag cgtgatgaat ttgtggtaaa gatagaagac atagagactt ttaaggaggc 2340
tttaaaaaca ggaaaagaac ctccagctat ttggaaagta caaaaagctt tattacagaa 2400
88/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
atttgttcct gaaattcgag atggtcaaag agaatttgct gctacaaata gttatcttgg 2460
atattttgga gatgcaaaga gtaaatacaa aagaatatat gtgaagttca ttgaaaatgc 2520
aaacaagaag gaatatgtca gagtgtgttc taaaaagcca agaaataaac cttcacaaac 2580
tatcagaact gttcaagcta agccaagtag tagcagtaaa acttctgatc ctctagcatc 2640
aaaaactaca actacaaaag ccccttccgt gaaacccaaa gttaaacagc caaaagtaaa 2700
ggctgagcca ccaccaaaga aacggaaaaa atggaaagaa gaattttcat catcccaatc 2760
tgactcatct cctgagatcc atactagtag tagtgacgat gaggaatttg aacctcccgc 2820
tccctttgtc actcgctttt tgaacacaag agcaatgaag gaaaccttta agagctacat 2880
ggaattgctt gttagcattg ccttggaccc tgacacaatg caagccttag agaagagcaa 2940
tgatgagcta cttttacctc atatgaaaaa aatagatggc atgctaaatg ataaccgaaa 3000
gagacttctt ttgaatcttc atttggatca atcattcaag aatgctttgg aaagttttcc 3060
tgaactaaca ataattactc gagattctaa agcaaagagt ggaggaactg ctatttctaa 3120
aatcaaaatg aatggcaaag cctataataa gaaaactcta aggacttcta aaacaaccac 3180
caaatctgca caagagtttg ctgtcgatcc agagaaaata cagttgtatt ctttgtatca 3240
ttcactccat cattataagt accatgttta tctgatatgt aaggatgaga tttcttcggt 3300
gcagaaaaaa aatgaagatt taggacagga ggaaattgtt caactttgta tgaaaaatgt 3360
aaaatgggtg gaggacctct ttgaaaaatt tggagaactt ctaaatcatg tacagcagaa 3420
atgttcctga cttttccaca aaaatcccat ctttttatag cactaatgaa atggcagata 3480
tggggtggtc aaagataatc agatgtcaag tagtggcctt ctgcaggccg gccgcttcca 3540
tcatggaact gtcattacca cctctgctga aggacagtgg tgcggccttt aggaacgaag 3600
ttagtcctct ggaaatggac ctaaatccca ccacattttt accctaatga atgatttttc 3660
tattttgtaa accattgggt aacttgagtc atattttcag aaacattttt tgacaaatga 3720
tgaagcatgc actaagtata attttttttt attgctagag aagtaacact taaagtaacg 3780
attttttttt tctgactccg gctaaacacc agaatgacag agaagtggca gaaaccatat 3840
gtttgtactc acatctggcc acaaaaccag aaatactgta cattatgtaa agaggtctgg 3900
tgtggtgtga catcctgtat aagaatatca tcaatttaaa atataaaatt tggaaactat 3960
tctgctttac agactccttt tactcttaac atgttcagga aactggatgt ggaattggtg 4020
caattctctg actgcttttt gtgtcaaatt atattgtgat aaaaaaacaa tgacatacta 4080
ttttccctat cgcaaagaaa agtattttcg tttatactgt tttttccttt ggaaaatttt 4140
caattgtaca ttttatttca ctgatagttg tatttttcac aaggaaaatg ttgtggttat 4200
aattacgttt gatatatctc tacaacacct tttgttattt tcagtaaatc ttagttatat 4260
gttgaatttc taatgtgaat tctatcttga ggtaaccatt tttttcatac agatttgctt 4320
cagtgttatc cagaatatgc attcagtact agaattagtt tagctttata aatagggctg 4380
tgttagacac tgcagtaatt ttctaattca taaaataaac ttcttactaa actagcactt 4440
gattaacttg ttgaggtaaa aatctaacta catttacatt ttgaagaata aaactgataa 4500
ttagactatt tgcagtgtta aacacagctt ccttaactct tagaactgga agttgtagag 4560
ctctcctttt ggtgcctttc cagcctttat acacactatt gtagctttct taggtttgat 4620
aggtagcgtt tcaagtagtt tagctgagac agtgaatgta ttaggttcaa catgaccttg 4680
tgttttattt gtgtttgcca acaggatgcc ttatttgttt gagaaaaaga tgtactagtg 4740
tcattctaaa ctatctcctt ttttaggatt ctaaagaagt to 4782
<210> 96
<211> 1410
<212> DNA
<213> Homo sapiens
<220>
<221> misc_feature
<223> Incyte ID No: 90089609CB1
<400> 96
aagaagtctg agcctgcctt agctgccccc caaggccatc caagctgtca ccgccaccac 60
ctccagcccc cctccctgcc atcctgcttc ccaggcagcc ctcactacca tccggcgacc 120
actctaagtc caagcatcca gcctccctgt cctaaccagc cacccccatc ggcccccata 180
cagcactaac caggagcatc ttagcccagc tgcctccctc ccgcccacga agctgtcctg 240
ccttcctggt gtcctggaac agagggaggc cctaacctgc ccattccagc tcccagagac 300
ataccctcct ctgggttcct tcacacccaa ccccctcttc tccttgaccc cacccatctc 360
cacccaccag tccctcccct tccttcaggg ctggggactc gtgggtgcag ctggcaccct 420
agtcctgcct gaccccccaa ggccaagggg gtacgtggaa ggggttacag aagcaagcag 480
atgaggcgca gcccagcccc cccggcacct tggtgagcgc cccggggtgg aaggtccagc 540
gcgtctcgtg gttgaactgc acgcgcacgt ccccgcggtc cgtgatacga tgcacggtgc 600
ccgtctgtcc gataaactgt ggcgggtcat ggagctgtgg ctgggtcggg ctggacgctt 660
ggggaggaag gggctgaggg ggcggagggc tgggagggca gggcacacag ggctccacgg 720
89/90
CA 02460480 2004-03-15
WO 03/031595 PCT/US02/32852
cggggcggct cacctccgcc atcctggggt tccagccgcc gtggccttcc tgcatctccc 780
gcaggacatc agtgtccagc agacacttga ccttgtcccc gtgctggaag ggctggctgt 840
cagcactcac cctgcgctgc agctccgccg gcttgcctgc ggagggtggc agaccaggcc 900
tcaggcagcg gcccctcccc tctgggactt cagccaaggc aggagggctg atggagtcag 960
tgtgacagcc tcataccgag ccttgggagg tggtccttgt agtagaagcc gcccgctgcc 1020
tcgcccacac acttgaggtc caccttgccc ttgtggccca cacggtacac attggtggta 1080
ccatcagccc acgtcacgct ggccacactc cggcctgtct ccacatccca gccacggatg 1140
tccaccacac ggcccggttt cccttcccct cctggggaaa gaggcccccc cagcaggcta 1200
aggctgagcc atgagccagc cacagcccag ccctgacccc gcccctctgc ctccactcac 1260
catcctgtga gccccactcc cagtcggggc ctcgcaccac cttcgctccc tggaagatgc 1320
cccttagtgg gatcctcggg aggccctggc ggggactcag tgtgacactg caagagggtt 1380
cacaaataaa gctctcagaa gaaaaaaaaa 1410
90/90
